bluuebunny/crossref_metadata_2025 huggingface dataset:
Glycine-Rich Proteins Encoded by a Nodule-Specific Gene Family Are Implicated in Different Stages of Symbiotic Nodule Development in Medicago Spp.
Authors: Kevei Zoltán , Vinardell José María , Kiss György B. , Kondorosi Adam , Kondorosi Eva
Published: May 2007

Four genes encoding small proteins with significantly high glycine content have been identified from root nodules of Medicago sativa. All of these proteins as well as their Medicago truncatula homologues carried an amino terminal signal peptide and a glycine-rich carboxy terminal domain. All except nodGRP3 lacked the characteristic repeat structure described for cell wall and stress response-related glycinerich proteins (GRP). Expression of these GRP genes was undetectable in flower, leaf, stem, and hypocotyl cells, whereas expression was highly induced during root nodule development, suggesting that GRP genes act as nodulins. Moreover, none of these nodule-expressed GRP genes were activated by hormones or stress treatments, which are inducers of many other GRPs. In Rhizobium-free spontaneous nodules and in nodules induced by a noninfective mutant strain of Sinorhizobium meliloti, all these genes were repressed, while they were induced in Fix¯ nodules, unaffected in bacterial infection, but halted in bacteroid differentiation. These results demonstrated that bacterial infection but not bacteroid differentiation is required for the induction of the nodule-specific GRP genes. Differences in kinetics and localization of gene activation as well as in the primary structure of proteins suggest nonredundant roles for these GRPs in nodule organogenesis.
Differential Virulence by Strains of Cucumber mosaic virus is Mediated by the 2b Gene
Authors: Shi Bu-Jun , Palukaitis Peter , Symons Robert H.
Published: May 2007

The approximately 12-kDa 2b protein, encoded by all cucumoviruses, had been shown to play an important role in viral long-distance movement, hypervirulence, and suppression of post-transcriptional gene silencing. The role of the 2b gene in the hypervirulence of Cucumber mosaic virus (CMV) and whether hypervirulence was linked to movement were analyzed using a hybrid virus (CMV-qw), generated by replacing the 2b gene in a subgroup II strain, Q-CMV, with the 2b gene from a subgroup IA strain, WAII-CMV. CMV-qw was more virulent than Q-CMV or WAII-CMV on most of the host plant species tested. Northern blot and nucleotide sequence analyses demonstrated that CMV-qw was stably maintained during the course of infection and upon passage. Kinetic studies revealed that the hypervirulence induced by the hybrid virus was associated with neither increased viral RNA accumulation nor more rapid viral movement per se, suggesting that other functions of the 2b protein are important in determining the hypervirulence.
A Pseudomonas syringae pv. tomato DC3000 Hrp (Type III Secretion) Deletion Mutant Expressing the Hrp System of Bean Pathogen P. syringae pv. syringae 61 Retains Normal Host Specificity for Tomato
Authors: Fouts Derrick E. , Badel Jorge L. , Ramos Adela R. , Rapp Ryan A. , Collmer Alan
Published: May 2007

The plant pathogenic species Pseudomonas syringae is divided into numerous pathovars based on host specificity. For example, P. syringae pv. tomato DC3000 is pathogenic on tomato and Arabidopsis, whereas P. syringae pv. syringae 61 is pathogenic on bean. The ability of P. syringae strains to elicit the hypersensitive response (HR) in non-hosts or be pathogenic (or parasitic) in hosts is dependent on the Hrp (type III secretion) system and effector proteins this system is thought to inject into plant cells. To test the role of the Hrp system in determining host range, the hrp/hrc gene cluster (hrpK through hrpR) was deleted from DC3000 and complemented in trans with the orthologous cluster from strain 61. Mutant CUCPB5114 expressing the bean pathogen Hrp system on plasmid pCPP2071 retained the ability of wild-type DC3000 to elicit the HR in bean, to grow and cause bacterial speck in tomato, and to elicit a cultivar-specific (gene-for-gene) HR in tomato plants carrying the Pto resistance gene. However, the symptoms produced in compatible tomato plants involved markedly reduced chlorosis, and CUCPB5114(pCPP2071) did not grow or produce symptoms in Arabidopsis Col-0 although it was weakly virulent in NahG Arabidopsis. A hypersensitive-like collapse was produced by CUCPB5114(pCPP2071) in Arabidopsis Col-0 at 1 × 107 CFU/ml, but only if the bacteria also expressed AvrB, which is recognized by the RPM1 resistance gene in Col-0 and confers incompatibility. These observations support the concept that the P. syringae effector proteins, rather than secretion system components, are the primary determinants of host range at both the species and cultivar levels of host specificity.
Induced Systemic Resistance in Arabidopsis thaliana in Response to Root Inoculation with Pseudomonas fluorescens CHA0
Authors: Iavicoli Annalisa , Boutet Emmanuel , Buchala Antony , Métraux Jean-Pierre
Published: May 2007

Root inoculation of Arabidopsis thaliana ecotype Columbia with Pseudomonas fluorescens CHA0r partially protected leaves from the oomycete Peronospora parasitica. The molecular determinants of Pseudomonas fluorescens CHA0r for this induced systemic resistance (ISR) were investigated, using mutants derived from strain CHA0: CHA400 (pyoverdine deficient), CHA805 (exoprotease deficient), CHA77 (HCN deficient), CHA660 (pyoluteorin deficient), CHA631 (2,4-diacetylphloroglucinol [DAPG] deficient), and CHA89 (HCN, DAPG- and pyoluteorin deficient). Only mutations interfering with DAPG production led to a significant decrease in ISR to Peronospora parasitica. Thus, DAPG production in Pseudomonas fluorescens is required for the induction of ISR to Peronospora parasitica. DAPG is known for its antibiotic activity; however, our data indicate that one action of DAPG could be due to an effect on the physiology of the plant. DAPG at 10 to 100 μM applied to roots of Arabidopsis mimicked the ISR effect. CHA0r-mediated ISR was also tested in various Arabidopsis mutants and transgenic plants: NahG (transgenic line degrading salicylic acid [SA]), sid2-1 (nonproducing SA), npr1-1 (non-expressing NPR1 protein), jar1-1 (insensitive to jasmonic acid and methyl jasmonic acid), ein2-1 (insensitive to ethylene), etr1-1 (insensitive to ethylene), eir1-1 (insensitive to ethylene in roots), and pad2-1 (phytoalexin deficient). Only jar1-1, eir1-1, and npr1-1 mutants were unable to undergo ISR. Sensitivity to jasmonic acid and functional NPR1 and EIR1 proteins were required for full expression of CHA0r-mediated ISR. The requirements for ISR observed in this study in Peronospora parasitica induced by Pseudomonas fluorescens CHA0r only partially overlap with those published so far for Peronospora parasitica, indicating a great degree of flexibility in the molecular processes leading to ISR.
Analysis of Gene Expression in Two Growth States ofXylella fastidiosaand Its Relationship with Pathogenicity
Authors: de Souza Alessandra A. , Takita Marco A. , Coletta-Filho Helvécio D. , Caldana Camila , Goldman Gustavo H. , Yanai Giane M. , Muto Nair H. , de Oliveira Regina C. , Nunes Luiz R. , Machado Marcos A.
Published: May 2007

Xylella fastidiosa is a plant pathogen responsible for diseases of economically important crops. Although there is considerable disagreement about its mechanism of pathogenicity, blockage of the vessels is one of the most accepted hypotheses. Loss of virulence by this bacterium was observed after serial passages in axenic culture. To confirm the loss of pathogenicity of X. fastidiosa, the causing agent of citrus variegated chlorosis (CVC), freshly-isolated bacteria (first passage [FP]condition) as well as bacteria obtained after 46 passages in axenic culture (several passage [SP]condition) were inoculated into sweet orange and periwinkle plants. Using real time quantitative polymerase chain reaction, we verified that the colonization of FP cells was more efficient for both hosts. The sequence of the complete X. fastidiosa genome allowed the construction of a DNA microarray that was used to investigate the total changes in gene expression associated with the FP condition. Most genes found to be induced in the FP condition were associated with adhesion and probably with adaptation to the host environment. This report represents the first study of the transcriptome of this pathogen, which has recently gained more importance, since the genome of several strains has been either partially or entirely sequenced.
Identical Accumulation and Immobilization of Sulfated and Nonsulfated Nod Factors in Host and Nonhost Root Hair Cell Walls
Authors: Goedhart Joachim , Bono Jean-Jacques , Bisseling Ton , Gadella Theodorus W. J.
Published: May 2007

Nod factors are signaling molecules secreted by Rhizobium bacteria. These lipo-chitooligosaccharides (LCOs) are required for symbiosis with legumes and can elicit specific responses at subnanomolar concentrations on a compatible host. How plants perceive LCOs is unclear. In this study, using fluorescent Nod factor analogs, we investigated whether sulfated and nonsulfated Nod factors were bound and perceived differently by Medicago truncatula and Vicia sativa root hairs. The bioactivity of three novel sulfated fluorescent LCOs was tested in a root hair deformation assay on M. truncatula, showing bioactivity down to 0.1 to 1 nM. Fluorescence microscopy of plasmolyzed M. truncatula root hairs shows that sulfated fluorescent Nod factors accumulate in the cell wall of root hairs, whereas they are absent from the plasma membrane when applied at 10 nM. When the fluorescent Nod factor distribution in medium surrounding a root was studied, a sharp decrease in fluorescence close to the root hairs was observed, visualizing the remarkable capacity of root hairs to absorb Nod factors from the medium. Fluorescence correlation microscopy was used to study in detail the mobilities of sulfated and nonsulfated fluorescent Nod factors which are biologically active on M. truncatula and V. sativa, respectively. Remarkably, no difference between sulfated and nonsulfated Nod factors was observed: both hardly diffuse and strongly accumulate in root hair cell walls of both M. truncatula and V. sativa. The implications for the mode of Nod factor perception are discussed.
Biocontrol Traits of Pseudomonas spp. Are Regulated by Phase Variation
Authors: van den Broek Daan , Chin-A-Woeng Thomas F. C. , Eijkemans Kevin , Mulders Ine H. M. , Bloemberg Guido V. , Lugtenberg Ben J. J.
Published: May 2007

Of 214 Pseudomonas strains isolated from maize rhizosphere, 46 turned out to be antagonistic, of which 43 displayed clear colony phase variation. The latter strains formed both opaque and translucent colonies, designated as phase I and phase II, respectively. It appeared that important biocontrol traits, such as motility and the production of antifungal metabolites, proteases, lipases, chitinases, and biosurfactants, are correlated with phase I morphology and are absent in bacteria with phase II morphology. From a Tn5luxAB transposon library of Pseudomonas sp. strain PCL1171 phase I cells, two mutants exhibiting stable expression of phase II had insertions in gacS. A third mutant, which showed an increased colony phase variation frequency was mutated in mutS. Inoculation of wheat seeds with PCL1171 bacteria of phase I morphology resulted in efficient suppression of take-all disease, whereas disease suppression was absent with phase II bacteria. Neither the gacS nor the mutS mutant was able to suppress take-all, but biocontrol activity was restored after genetic complementation of these mutants. Furthermore, in a number of cases, complementation by gacS of wild-type phase II sectors to phase I phenotype could be shown. A PCL1171 phase I mutant defective in antagonistic activity appeared to have a mutation in a gene encoding a lipopeptide synthetase homologue and had lost its biocontrol activity, suggesting that biocontrol by strain PCL1171 is dependent on the production of a lipopeptide. Our results show that colony phase variation plays a regulatory role in biocontrol by Pseudomonas bacteria by influencing the expression of major biocontrol traits and that the gacS and mutS genes play a role in the colony phase variation process. Therefore phase variation not only plays a role in escaping animal defense but it also appears to play a much broader and vital role in the ecology of bacteria producing exoenzymes, antibiotics, and other secondary metabolites.
Interactions in the Tomato Rhizosphere of TwoPseudomonasBiocontrol Strains with the Phytopathogenic FungusFusarium oxysporumf. sp.radicis-lycopersici
Authors: Bolwerk Annouschka , Lagopodi Anastasia L. , Wijfjes André H. M. , Lamers Gerda E. M. , Chin-A-Woeng Thomas F. C. , Lugtenberg Ben J. J. , Bloemberg Guido V.
Published: May 2007

The fungus Fusarium oxysporum f. sp. radicis-lycopersici causes foot and root rot of tomato plants, which can be controlled by the bacteria Pseudomonas fluorescens WCS365 and P. chlororaphis PCL1391. Induced systemic resistance is thought to be involved in biocontrol by P. fluorescens WCS365. The antifungal metabolite phenazine-1-carbox-amide (PCN), as well as efficient root colonization, are essential in the mechanism of biocontrol by P. chlororaphis PCL1391. To understand the effects of bacterial strains WCS365 and PCL1391 on the fungus in the tomato rhizosphere, microscopic analyses were performed using different autofluorescent proteins as markers. Tomato seedlings were inoculated with biocontrol bacteria and planted in an F. oxysporum f. sp. radicis-lycopersici-infested gnotobiotic sand system. Confocal laser scanning microscope analyses of the interactions in the tomato rhizosphere revealed that i) the microbes effectively compete for the same niche, and presumably also for root exudate nutrients; ii) the presence of either of the two bacteria negatively affects infection of the tomato root by the fungus; iii) both biocontrol bacteria colonize the hyphae extensively, which may represent a new mechanism in biocontrol by these pseudomonads; and iv) the production of PCN by P. chlororaphis PCL1391 negatively affects hyphal growth and branching, which presumably affects the colonization and infecting ability of the fungus.
Three Genes Encoding for Putative Methyl- and Acetyltransferases Map Adjacent to the wzm and wzt Genes and Are Essential for O-Antigen Biosynthesis in Rhizobium etli CE3
Authors: Lerouge I. , Verreth C. , Michiels J. , Carlson R. W. , Datta A. , Gao M.-Y. , Vanderleyden J.
Published: May 2007

The elucidation of the structure of the O-antigen of Rhizo-bium etli CE3 predicts that the R. etli CE3 genome must contain genes encoding acetyl- and methyltransferases to confer the corresponding modifications to the O-antigen. We identified three open reading frames (ORFs) upstream of wzm, encoding the membrane component of the O-antigen transporter and located in the lpsα-region of R. etli CE3. The ORFs encode two putative acetyltransferases with similarity to the CysE-LacA-LpxA-NodL family of acetyl-transferases and one putative methyltransferase with sequence motifs common to a wide range of S-adenosyl-L-methionine-dependent methyltransferases. Mutational analysis of the ORFs encoding the putative acetyltrans-ferases and methyltransferase revealed that the acetyl and methyl decorations mediated by these specific enzymes are essential for O-antigen synthesis. Composition analysis and high performance anion exchange chromatography analysis of the lipopolysaccharides (LPSs) of the mutants show that all of these LPSs contain an intact core region and lack the O-antigen polysaccharide. The possible role of these transferases in the decoration of the O-antigen of R. etli is discussed.
Laminarin Elicits Defense Responses in Grapevine and Induces Protection AgainstBotrytis cinereaandPlasmopara viticola
Authors: Aziz Aziz , Poinssot Benoît , Daire Xavier , Adrian Marielle , Bézier Annie , Lambert B. , Joubert Jean-Marie , Pugin Alain
Published: May 2007

Grapevine (Vitis vinifera L.) is susceptible to many pathogens, such as Botrytis cinerea, Plasmopara viticola, Uncinula necator, and Eutypa lata. Phytochemicals are used intensively in vineyards to limit pathogen infections, but the appearance of pesticide-resistant pathogen strains and a desire to protect the environment require that alternative strategies be found. In the present study, the β-1,3-glucan laminarin derived from the brown algae Laminaria digitata was shown both to be an efficient elicitor of defense responses in grapevine cells and plants and to effectively reduce B. cinerea and P. viticola development on infected grapevine plants. Defense reactions elicited by laminarin in grapevine cells include calcium influx, alkalinization of the extracellular medium, an oxidative burst, activation of two mitogen-activated protein kinases, expression of 10 defense-related genes with different kinetics and intensities, increases in chitinase and β-1,3-glucanase activities, and the production of two phytoalexins (resveratrol and ε-viniferin). Several of these effects were checked and confirmed in whole plants. Laminarin did not induce cell death. When applied to grapevine plants, laminarin reduced infection by B. cinerea and P. viticola by approximately 55 and 75%, respectively. Our data describing a large set of defense reactions in grapevine indicate that the activation of defense responses using elicitors could be a valuable strategy to protect plants against pathogens.
QTLs for Tomato Powdery Mildew Resistance (Oidium lycopersici) in Lycopersicon parviflorum G1.1601 Co-localize with Two Qualitative Powdery Mildew Resistance Genes
Authors: Bai Yuling , Huang Cai-Cheng , van der Hulst Ron , Meijer-Dekens Fien , Bonnema Guusje , Lindhout Pim
Published: May 2007

Tomato (Lycopersicon esculentum) is susceptible to the powdery mildew Oidium lycopersici, but several wild relatives such as Lycopersicon parviflorum G1.1601 are completely resistant. An F2 population from a cross of Lycopersicon esculentum cv. Moneymaker × Lycopersicon parviflorum G1.1601 was used to map the O. lycopersici resistance by using amplified fragment length polymorphism markers. The resistance was controlled by three quantitative trait loci (QTLs). Ol-qtl1 is on chromosome 6 in the same region as the Ol-1 locus, which is involved in a hypersensitive resistance response to O. lycopersici. Ol-qtl2 and Ol-qtl3 are located on chromosome 12, separated by 25 cM, in the vicinity of the Lv locus conferring resistance to another powdery mildew species, Leveillula taurica. The three QTLs, jointly explaining 68% of the phenotypic variation, were confirmed by testing F3 progenies. A set of polymerase chain reaction-based cleaved amplified polymorphic sequence and sequence characterized amplified region markers was generated for efficient monitoring of the target QTL genomic regions in marker assisted selection. The possible relationship between genes underlying major and partial resistance for tomato powdery mildew is discussed.
Expression of the Bacterial Catalase Genes During Sinorhizobium meliloti-Medicago sativa Symbiosis and Their Crucial Role During the Infection Process
Authors: Jamet Alexandre , Sigaud Samuel , Van de Sype Ghislaine , Puppo Alain , Hérouart Didier
Published: May 2007

Sinorhizobium meliloti possesses three distinct catalases to cope with oxidative stress: two monofunctional catalases (KatA and KatC) and one bifunctional catalase-peroxydase (KatB). The katB gene is constitutively expressed during growth in batch culture and is not induced under oxidative stress conditions. In contrast, the expression of katA and katC genes is mainly regulated at the transcription level in these conditions. A differential expression of kat genes was observed during the development of the nodule. A high expression of katA gene was detected in bacteroids, suggesting that the nitrogen-fixation process induces a strong oxidative stress. In contrast, bacteria express katB and katC genes and not the H2O2-inducible katA gene in infection threads despite the detection of H2O2 around the bacteria. A katB katC double mutant nodulated poorly and displayed abnormal infection. After nonefficient release into plant cells, bacteria failed to differentiate into bacteroids and rapidly underwent senescence. Our results indicate that these two catalases are essential for the establishment of the symbiosis. They also suggest that the bacteria are in a nonexponential growth phase in infection threads and corroborate previous studies on the growth rate of bacteria inside the plant.
Members of the Arabidopsis WRKY Group III Transcription Factors Are Part of Different Plant Defense Signaling Pathways
Authors: Kalde Monika , Barth Meike , Somssich Imre E. , Lippok Bernadette
Published: May 2007

WRKY proteins are a large group of transcription factors restricted to the plant kingdom. In Arabidopsis thaliana, the gene family consists of 74 members. Here, we analyzed the expression of all 13 members of one main WRKY subgroup and found that the majority are responsive both to pathogen infection and to salicylic acid. Temporal expression studies during compatible, incompatible, and nonhost interactions and employing plant defense-signaling mutants allowed us to define four distinct WRKY subsets responding to different signaling queues along defense pathways. These subsets did not reflect phylogenetic relationships. Promoter studies of one member, AtWRKY54, using a reporter gene construct in transgenic Arabidopsis plants, revealed that regulatory regions mediating pathogen and SA inducibility are clearly separable. In an AtWRKY54 knockout line, resistance to Peronospora parasitica was not compromised, but the transient expression kinetics of several WRKY genes was affected, suggesting both the existence of functional redundancy and intense cross-talk between signaling networks.
The Colletotrichum lagenarium Ste12-Like Gene CST1 Is Essential for Appressorium Penetration
Authors: Tsuji Gento , Fujii Satoshi , Tsuge Seiji , Shiraishi Tomonori , Kubo Yasuyuki
Published: May 2007

Colletotrichum lagenarium is the causal agent of anthracnose of cucumber. This fungus produces a darkly melanized infection structure, appressoria, to penetrate the host leaves. The C. lagenarium CMK1 gene, a homologue of the Saccharomyces cerevisiae FUS3/KSS1 mitogen-activated protein (MAP) kinase genes, was shown to regulate conidial germination, appressorium formation, and invasive growth. In S. cerevisiae, Ste12p is known to be a transcriptional factor downstream of Fus3p/Kss1p MAP kinases. To evaluate the CMK1 MAP kinase pathway, we isolated the Ste12 homologue CST1 gene from C. lagenarium and characterized. The cst1Δ strains were nonpathogenic on intact host leaves, but could form lesions when inoculated on wounded leaves. Conidia of the cst1Δ strains could germinate and form melanized appressoria on both host leaf surface and artificial cellulose membrane, but could not produce infectious hyphae from appressoria, suggesting that CST1 is essential for appressorium penetration in C. lagenarium. In addition, matured appressoria of the cst1Δ strains contained an extremely low level of lipid droplets compared with that of the wild-type strain. Lipid droplets were abundant in conidia of the cst1Δ strains, but rapidly disappeared during appressorium formation. This misscheduled lipid degradation might be related to the failure of appressorium penetration in the cst1Δ strain.
New NodW- or NifA-Regulated Bradyrhizobium japonicum Genes
Authors: Baumberger Isabelle Caldelari , Fraefel Nicole , Göttfert Michael , Hennecke Hauke
Published: May 2007

A cluster of genes coding for putative plant cell-wall degrading enzymes (i.e., genes for two endoglucanases [gunA and gunA2], one pectinmethylesterase [pme], and one polygalacturonase [pgl]) was identified by sequence similarities in the symbiotic region of the Bradyrhizobium japonicum chromosome. In addition, a systematic screen of the region revealed several genes potentially transcribed by the σ54-RNA polymerase and activated by the transcriptional regulator NifA (i.e., genes for proteins with similarity to outer membrane proteins [id117 and id525] and a citrate carrier [id331 or citA] and one open reading frame without similarity to known proteins [id747]). Expression studies using transcriptional lacZ fusions showed that gunA2 and pgl were strongly induced by the isoflavone genistein in a NodW-dependent manner, suggesting a role of the gene products in early events of the nodulation process; by contrast, gunA and pme expression was very weak in the conditions tested. The gunA2 gene product was purified and was shown to have cellulase activity. β-Galactosidase activity expressed from transcriptional lacZ fusions to id117, id525, and id747 in the wild type and in nifA and rpoN mutant backgrounds confirmed that their transcription was dependent on NifA and σ54. Despite the presence of a -24/-12-type promoter and a NifA binding site upstream of citA, no regulation could be demonstrated in this case. Null mutations introduced in gunA, gunA2, pgl, pme, citA, id117, id525, and id747 did not impair the symbiosis with the host plants.
Characterization of a Novel Barley Protein, HCP1, That Interacts with theBrome mosaic virusCoat Protein
Authors: Okinaka Yasushi , Mise Kazuyuki , Okuno Tetsuro , Furusawa Iwao
Published: May 2007

Brome mosaic virus (BMV) requires the coat protein (CP) not only for encapsidation but also for viral cell-to-cell and long-distance movement in barley plants. This suggests that BMV infection is controlled by interactions of CP with putative host factors as well as with viral components. To identify the host factors that interact with BMV CP, we screened a barley cDNA library containing 2.4 × 106independent clones, using a yeast two-hybrid system. Using full-length and truncated BMV CPs as baits, four candidate cDNA clones were isolated. One of the candidate cDNAs encodes a unique oxidoreductase enzyme, designated HCP1. HCP1 was found predominantly in the soluble fractions after differential centrifugation of BMV-infected and mock-inoculated barley tissues. A two-hybrid binding assay using a series of truncated BMV CPs demonstrated that a C-terminal portion of CP is essential for its interaction with HCP1. Interestingly, experiments with CP mutants bearing single amino acid substitutions at the C-terminus revealed that the capacity for mutant CP-HCP1 binding correlates well with the infectivity of the corresponding mutant viruses in barley. These results indicate that CP-HCP1 binding controls BMV infection of barley, interacting directly with CP, probably in the cell cytoplasm.
A Profile of Putative Parasitism Genes Expressed in the Esophageal Gland Cells of the Root-knot Nematode Meloidogyne incognita
Authors: Huang Guozhong , Gao Bingli , Maier Tom , Allen R. , Davis Eric L. , Baum Thomas J. , Hussey Richard S.
Published: May 2007

Identifying parasitism genes encoding proteins secreted from a nematode's esophageal gland cells and injected through its stylet into plant tissue is the key to understanding the molecular basis of nematode parasitism of plants. Meloidogyne incognita parasitism genes were cloned by microaspirating the cytoplasm from the esophageal gland cells of different parasitic stages to provide mRNA to create a gland cell-specific cDNA library by long-distance reverse-transcriptase polymerase chain reaction. Of 2,452 cDNA clones sequenced, deduced protein sequences of 185 cDNAs had a signal peptide for secretion and, thus, could have a role in root-knot nematode parasitism of plants. High-throughput in situ hybridization with cDNA clones encoding signal peptides resulted in probes of 37 unique clones specifically hybridizing to transcripts accumulating within the subventral (13 clones) or dorsal (24 clones) esophageal gland cells of M. incognita. In BLASTP analyses, 73% of the predicted proteins were novel proteins. Those with similarities to known proteins included a pectate lyase, acid phosphatase, and hypothetical proteins from other organisms. Our cell-specific analysis of genes encoding secretory proteins provided, for the first time, a profile of putative parasitism genes expressed in the M. incognita esophageal gland cells throughout the parasitic cycle.
Peh Production, Flagellum Synthesis, and Virulence Reduced in Erwinia carotovora subsp. carotovora by Mutation in a Homologue of cytR
Authors: Matsumoto Hiroyuki , Muroi Hironobu , Umehara Masahiro , Yoshitake Yoshimasa , Tsuyumu Shinji
Published: May 2007

Erwinia carotovora subsp. carotovora is a causal agent of soft-rot diseases in a wide variety of plants. Here, we have isolated a new regulatory factor involved in the virulence of E. carotovora subsp. carotovora by in vivo insertional mutagenesis using a transposon Tn5. The gene was homologous to cytR encoding a transcriptional repressor of nucleoside uptake and catabolism genes in Escherichia coli, Salmonella typhimurium, and Vibrio cholerae. Phenotypic characterization of a nonpolar deletion mutant of the cytR homologue (ΔcytR) revealed that the ΔcytR mutant produced a reduced level of polygalacturonase (Peh) and lost its motility compared to that in the parental strain. With electron microscopy, the ΔcytR mutant was shown to be aflagellate. Furthermore, the expression of fliA and fliC (encoding σ28 and flagellin, respectively) was also reduced in ΔcytR mutant. The virulence of ΔcytR mutant was reduced in Chinese cabbage and potato compared to that of the parental strain. These results suggest that the CytR homologue of E. carotovora subsp. carotovora positively controls Peh production and flagellum synthesis and plays an important role in its pathogenicity.
Two Potato Proteins, Including a Novel RING Finger Protein (HIP1), Interact with the Potyviral Multifunctional Protein HCpro
Authors: Guo Deyin , Spetz Carl , Saarma Mart , Valkonen Jari P. T.
Published: May 2007

Potyviral helper-component proteinase (HCpro) is a multifunctional protein exerting its cellular functions in interaction with putative host proteins. In this study, cellular protein partners of the HCpro encoded by Potato virus A (PVA) (genus Potyvirus) were screened in a potato leaf cDNA library using a yeast two-hybrid system. Two cellular proteins were obtained that interact specifically with PVA HCpro in yeast and in the two in vitro binding assays used. Both proteins are encoded by single-copy genes in the potato genome. Analysis of the deduced amino acid sequences revealed that one (HIP1) of the two HCpro interactors is a novel RING finger protein. The sequence of the other protein (HIP2) showed no resemblance to the protein sequences available from databanks and has known biological functions.
A Protein Kinase from Colletotrichum trifolii Is Induced by Plant Cutin and Is Required for Appressorium Formation
Authors: Dickman M. B. , Ha Y.-S. , Yang Z. , Adams B. , Huang C.
Published: May 2007

When certain phytopathogenic fungi contact plant surfaces, specialized infection structures (appressoria) are produced that facilitate penetration of the plant external barrier; the cuticle. Recognition of this hydrophobic host surface must be sensed by the fungus, initiating the appropriate signaling pathway or pathways for pathogenic development. Using polymerase chain reaction and primers designed from mammalian protein kinase C sequences (PKC), we have isolated, cloned, and characterized a protein kinase from Colletotrichum trifolii, causal agent of alfalfa anthracnose. Though sequence analysis indicated conserved sequences in mammalian PKC genes, we were unable to induce activity of the fungal protein using known activators of PKC. Instead, we show that the C. trifolii gene, designated LIPK (lipidinduced protein kinase) is induced specifically by purified plant cutin or long-chain fatty acids which are monomeric constituents of cutin. PKC inhibitors prevented appressorium formation and, to a lesser extent, spore germination. Overexpression of LIPK resulted in multiple, abnormally shaped appressoria. Gene replacement of lipk yielded strains which were unable to develop appressoria and were unable to infect intact host plant tissue. However, these mutants were able to colonize host tissue following artificial wounding, resulting in typical anthracnose lesions. Taken together, these data indicate a central role in triggering infection structure formation for this protein kinase, which is induced specifically by components of the plant cuticle. Thus, the fungus is able to sense and use host surface chemistry to induce a protein kinase-mediated pathway that is required for pathogenic development.
Transcriptional Silencing of Geminiviral Promoter-Driven Transgenes Following Homologous Virus Infection
Authors: Seemanpillai Mark , Dry Ian , Randles John , Rezaian Ali
Published: May 2007

Promoters isolated from the Tomato leaf curl virus (TLCV) drive both constitutive and tissue-specific expression in transgenic tobacco. Following systemic TLCV infection of plants stably expressing TLCV promoter:GUS transgenes, transgene expression driven by all six TLCV promoters was silenced. Silencing in the TLCV coat protein promoter:GUS plants (V2:GUSΔC) was characterized in more detail. Transgene silencing observed in leaf, stem, and preanthesis floral tissue occurred with the continued replication of TLCV in host tissues. Infection of the V2:GUSΔC plants with heterologous geminiviruses did not result in transgene silencing, indicating that silencing was specifically associated with TLCV infection. Nuclear run-on assays indicated that silencing was due to the abolition of transcription from the V2:GUSΔC transgene. Bisulfite sequencing showed that silencing was associated with cytosine hypermethylation of the TLCV-derived promoter sequences of the V2:GUSΔC transgene. Progeny derived from V2:GUSΔC plants silenced by TLCV infection were analyzed. Transgene expression was silenced in progeny seedlings but was partially reactivated in the majority of plants by 75 days postgermination. Progeny seedlings treated with the nonmethylatable cytosine analog 5-azacytidine or the histone deacetylase inhibitor sodium butyrate exhibited partial reactivation of expression. This is the first report of the hypermethylation of a virus-derived transgene associated with a DNA virus infection.
Novel Exchangeable Effector Loci Associated with the Pseudomonas syringae hrp Pathogenicity Island: Evidence for Integron-Like Assembly from Transposed Gene Cassettes
Authors: Charity James C. , Pak Kyong , Delwiche Charles F. , Hutcheson Steven W.
Published: May 2007

Pseudomonas syringae strains use a type III secretion system (TTSS) to translocate effector proteins that assist in the parasitism of host plant cells. Some genes for effector proteins are clustered in the exchangeable effector locus (EEL) associated with the hrp pathogenicity island. A polymerase chain reaction-based screen was developed to amplify the EEL from P. syringae strains. Of the 86 strains screened, the EEL was successfully amplified from 30 predominately North American P. syringae pv. syringae strains using hrpK and queA-derived primers and from an additional three strains using hrpL and queA-derived primers. Among the amplified EEL, ten distinct types of EEL were identified that could be classified into six families distinguishable by genetic composition, but other types of EEL may be present in strains isolated in other geographical regions. No linkage with the host range of the source strain was apparent. Gene cassettes carrying conserved flanking, coding, and intergenic sequences, present in different combinations, were identified in the characterized EEL. Six new alleles of known effectors were identified that differed from the homolog in sequence, size, or both of the gene. One of these apparently novel effector proteins, HopPsyB, retained a strongly conserved amino terminus similar to that of HopPsyA, but other regions of the two polypeptides were only weakly similar. hopPsyB was expressed from an apparent operon that included hrpK and a shcA homolog, shcB. Escherichia coli MC4100 expressing the hrp TTSS, ShcB, and HopPsyB elicited the hypersensitive response (HR) in tobacco, consistent with effector production. Indicative of translocation as an effector, P. syringae pv. tomato DC3000 expressing a HopPsyB':'AvrRpt2 fusion elicited the HR in RPS2+ Arabidopsis thaliana. P. syringae pv. tomato DC3000 carrying HopPsyB exhibited slightly enhanced virulence in several Brassica spp. These results are consistent with the hypotheses that the EEL is a source of disparate effectors functioning in pathogenicity of P. syringae strains and that it evolved independently of the hrp pathogenicity island central conserved region, most likely through integron-like assembly of transposed gene cassettes.
Multiple Resistance Phenotypes to Lettuce mosaic virus Among Arabidopsis thaliana Accessions
Authors: Revers Frédéric , Guiraud Thomas , Houvenaghel Marie-Christine , Mauduit Thierry , Le Gall Olivier , Candresse Thierry
Published: May 2007

With the aim to characterize plant and viral factors involved in the molecular interactions between plants and potyviruses, a Lettuce mosaic virus (LMV)-Arabidopsis thaliana pathosystem was developed. Screening of Arabi-dopsis accessions with LMV isolates indicated the existence of a large variability in the outcome of the interaction, allowing the classification of Arabidopsis accessions into seven susceptibility groups. Using a reverse genetic approach, the genome-linked protein of LMV, a multifunc-tional protein shown to be involved in viral genome amplification and movement of potyviruses, was established as the viral determinant responsible for the ability to overcome the resistance of the Niederzenz accession to LMV-0. Preliminary genetic analyses from F2 and recombinant inbred lines available between susceptible and resistant Arabidopsis accessions revealed the existence of at least three resistance phenotypes to LMV with different genetic bases. One dominant resistance gene, designated LLM1, involved in blocking the replication or cell-to-cell movement of the LMV-0 isolate in the Columbia accession, was mapped to chromosome I and shown to be linked to the marker nga280. At the same time, genetic analyses of segregating F2 populations were consistent with the restriction of the systemic movement of the LMV-AF199 isolate in Columbia being controlled by two dominant genes and with the complete resistance to all tested LMV isolates of the Cape Verde islands (Cvi) accession being conferred by a single recessive resistance gene. Sequencing of the eu-karyotic translation initiation factor 4E genes from the different LMV-resistant Arabidopsis accessions showed that these factors are not directly involved in the characterized resistance phenotypes.
The Dual Role of the Potyvirus P3 Protein ofTurnip mosaic virusas a Symptom and Avirulence Determinant in Brassicas
Authors: Jenner Carol E. , Wang Xiaowu , Tomimura Kenta , Ohshima Kazusato , Ponz Fernando , Walsh John A.
Published: May 2007

Two isolates of the potyvirus Turnip mosaic virus (TuMV), UK 1 and CDN 1, differ both in their general symptoms on the susceptible propagation host Brassica juncea and in their ability to infect B. napus lines possessing a variety of dominant resistance genes. The isolate CDN 1 produces a more extreme mosaic in infected brassica leaves than UK 1 and is able to overcome the resistance genes TuRB01, TuRB04, and TuRB05. The resistance gene TuRB03, in the B. napus line 22S, is effective against CDN 1 but not UK 1. The nucleic acid sequences of the UK 1 and CDN 1 isolates were 90% identical. The C-terminal half of the P3 protein was identified as being responsible for the differences in symptoms in B. juncea. A single amino acid in the P3 protein was found to be the avirulence determinant for TuRB03. Previous work already has identified the P3 as an avirulence determinant for TuRB04. Our results increase the understanding of the basis of plant-virus recognition, show the importance of the potyviral P3 gene as a symptom determinant, and provide a role in planta for the poorly understood P3 protein in a normal infection cycle.
Genetic and Physical Localization of the Soybean Rpg1-b Disease Resistance Gene Reveals a Complex Locus Containing Several Tightly Linked Families of NBS-LRR Genes
Authors: Ashfield Tom , Bocian Anna , Held Dan , Henk Adam D. , Marek Laura Fredrick , Danesh Dariush , Peñuela Silvia , Meksem Khalid , Lightfoot David A. , Young Nevin D. , Shoemaker Randy C. , Innes Roger W.
Published: May 2007

Alleles or tightly linked genes at the soybean (Glycine max L. Merr.) Rpg1 locus confer resistance to strains of Pseudomonas syringae pv. glycinea that express the avirulence genes avrB or avrRpm1. We have previously mapped Rpg1-b (the gene specific for avrB) to a cluster of resistance genes (R genes) with diverse specificities in molecular linkage group F. Here, we describe the high-resolution physical and genetic mapping of Rpg1-b to a 0.16-cM interval encompassed by two overlapping BAC clones spanning approximately 270 kilobases. Rpg1-b is part of a complex locus containing numerous genes related to previously characterized coiled coil-nucleotide binding site-leucine rich repeat (CC-NBS-LRR)-type R genes that are spread throughout this region. Phylogenetic and Southern blot analyses group these genes into four distinct subgroups, some of which are conserved in the common bean, Phaseolus vulgaris, indicating that this R gene cluster may predate the divergence of Phaseolus and Glycine. Members from different subgroups are physically intermixed and display a high level of polymorphism between soybean cultivars, suggesting that this region is rearranging at a high frequency. At least five CC-NBS-LRR-type genes cosegregate with Rpg1-b in our large mapping populations.
Rme1 is Necessary for Mi-1-Mediated Resistance and Acts Early in the Resistance Pathway
Authors: de Ilarduya Oscar Martinez , Nombela Gloria , Hwang Chin-Feng , Williamson Valerie M. , Muñiz Mariano , Kaloshian Isgouhi
Published: May 2007

The tomato gene Mi-1 confers resistance to root-knot nematodes (Meloidogyne spp.), potato aphid, and whitefly. Using genetic screens, we have isolated a mutant, rme1 (resistance to Meloidogyne spp.), compromised in resistance to M. javanica and potato aphid. Here, we show that the rme1 mutant is also compromised in resistance to M. incognita, M. arenaria, and whitefly. In addition, using an Agrobacterium-mediated transient assay in leaves to express constitutive gain-of-function mutant Pto L205D, we demonstrated that the rme1 mutation is not compromised in Pto-mediated hypersensitive response. Moreover, the mutation in rme1 does not result in increased virulence of pathogenic Pseudomonas syringae or Mi-1-virulent M. incognita. Using a chimeric Mi-1 construct, Mi-DS4, which confers constitutive cell death phenotype and A. rhizogenes root transformation, we showed that the Mi-1-mediated cell death pathway is intact in this mutant. Our results indicate that Rme1 is required for Mi-1-mediated resistance and acts either at the same step in the signal transduction pathway as Mi-1 or upstream of Mi-1.
Rhizoremediation: A Beneficial Plant-Microbe Interaction
Authors: Kuiper Irene , Lagendijk Ellen L. , Bloemberg Guido V. , Lugtenberg Ben J. J.
Published: May 2007

Worldwide, contamination of soil and ground water is a severe problem. The negative effects of pollutants on the environment and on human health are diverse and depend on the nature of the pollution. The search for alternative methods for excavation and incineration to clean polluted sites resulted in the application of bioremediation techniques. In this review, we describe some generally accepted bioremediation tools and subsequently focus on the combination of two approaches, phytoremediation and bioaugmentation, resulting in rhizoremediation. During rhizoremediation, exudates derived from the plant can help to stimulate the survival and action of bacteria, which subsequently results in a more efficient degradation of pollutants. The root system of plants can help to spread bacteria through soil and help to penetrate otherwise impermeable soil layers. The inoculation of pollutant-degrading bacteria on plant seed can be an important additive to improve the efficiency of phytoremediation or bioaugmentation.
cDNA Macroarray Analysis of Gene Expression in Ineffective Nodules Induced on the Lotus japonicus sen1 Mutant
Authors: Suganuma Norio , Yamamoto Atsuko , Itou Ai , Hakoyama Tsuneo , Banba Mari , Hata Shingo , Kawaguchi Masayoshi , Kouchi Hiroshi
Published: May 2007

The Lotus japonicus sen1 mutant forms ineffective nodules in which development is arrested at the stage of bacterial differentiation into nitrogen-fixing bacteroids. Here, we used cDNA macroarray systems to compare gene expression in ineffective nodules induced on the sen1 mutant with gene expression in wild-type nodules, in order to identify the host plant genes that are involved in nitrogen fixation. Macroarray analysis coupled with Northern blot analysis revealed that the expression of 18 genes was significantly enhanced in ineffective sen1 nodules, whereas the expression of 30 genes was repressed. Many of the enhanced genes encoded hydrolase enzymes, such as cysteine proteinase and asparaginase, that might function in the early senescence of sen1 nodules. By contrast, the repressed genes encoded nodulins, enzymes that are involved in carbon and nitrogen metabolism, membrane transporters, enzymes involved in phytohormone metabolism and secondary metabolism, and regulatory proteins. These proteins might have a role in the establishment of nitrogen fixation. In addition, we discovered two novel genes that encoded glutamate-rich proteins and were localized in the vascular bundles of the nodules. The expression of these genes was repressed in the ineffective nodules, which had lower levels of nitrogenase activity.
Involvement of N-acylhomoserine Lactones Throughout Plant Infection by Erwinia carotovora subsp. atroseptica (Pectobacterium atrosepticum)
Authors: Smadja Bruno , Latour Xavier , Faure Denis , Chevalier Sylvie , Dessaux Yves , Orange Nicole
Published: May 2007

Erwinia carotovora subsp. atroseptica is responsible for potato blackleg disease in the field and tuber soft rot during crop storage. The process leading to the disease occurs in two phases: a primary invasion step followed by a maceration step. Bacteria-to-bacteria communication is associated with a quorum-sensing (QS) process based on the production of N-acylhomoserine lactones (HSL). The role of HSL throughout plant infection was analyzed. To this purpose, HSL produced by a specific E. carotovora subsp. atroseptica wild-type strain, which was particularly virulent on potato, were identified. A derivative of this strain that expressed an HSL lactonase gene and produced low amounts of HSL was generated. The comparison of these strains allowed the evaluation of the role of HSL and QS in disease establishment and development. Bacterial growth and motility; activity of proteins secreted by type I, II, and III systems; and hypersensitive and maceration reactions were evaluated. Results indicated that HSL production and QS regulate only those traits involved in the second stage of the host plant infection (i.e., tissue maceration) and hypersensitive response in nonhost tobacco plants. Therefore, the use of QS quenching strategies for biological control in E. carotovora subsp. atroseptica cannot prevent initial infection and multiplication of this pathogen.
The Rice Bacterial Blight Resistance Gene xa5 Encodes a Novel Form of Disease Resistance
Authors: Iyer Anjali S. , McCouch Susan R.
Published: May 2007

The rice xa5 gene for disease resistance to Xanthomonas oryzae pv. oryzae has been positionally cloned and encodes the gamma subunit of transcription factor IIA (TFIIAγ). TFIIAγ is a general eukaryotic transcription factor with no previously known role in disease resistance. xa5 is unusual in that it is recessive and does not conform to one of the typical resistance gene structural classes. Sequencing of TFIIAγ in resistant and susceptible isolines revealed two nucleotide substitutions resulting in an amino acid change between resistant and susceptible cultivars. This association was conserved across 27 resistant and nine susceptible rice lines in the Aus-Boro group.