bluuebunny/crossref_metadata_2025 huggingface dataset:
New Nodulation Mutants Responsible for Infection Thread Development in Lotus japonicus
Authors: Yano Koji , Tansengco Myra L. , Hio Taihei , Higashi Kuniko , Murooka Yoshikatsu , Imaizumi-Anraku Haruko , Kawaguchi Masayoshi , Hayashi Makoto
Published: May 2007

Legume plants develop specialized root organs, the nodules, through a symbiotic interaction with rhizobia. The developmental process of nodulation is triggered by the bacterial microsymbiont but regulated systemically by the host legume plants. Using ethylmethane sulfonate mutagenesis as a tool to identify plant genes involved in symbiotic nodule development, we have isolated and analyzed five nodulation mutants, Ljsym74-3, Ljsym79-2, Ljsym79-3, Ljsym80, and Ljsym82, from the model legume Lotus japonicus. These mutants are defective in developing functional nodules and exhibit nitrogen starvation symptoms after inoculation with Mesorhizobium loti. Detailed observation revealed that infection thread development was aborted in these mutants and the nodules formed were devoid of infected cells. Mapping and complementation tests showed that Ljsym74-3, and Ljsym79-2 and Ljsym79-3, were allelic with reported mutants of L. japonicus, alb1 and crinkle, respectively. The Ljsym82 mutant is unique among the mutants because the infection thread was aborted early in its development. Ljsym74-3 and Ljsym80 were characterized as mutants with thick infection threads in short root hairs. Map-based cloning and molecular characterization of these genes will help us understand the genetic mechanism of infection thread development in L. japonicus.
Bacterial Endophytes and Their Interactions with Hosts
Authors: Rosenblueth Mónica , Martínez-Romero Esperanza
Published: May 2007

Recent molecular studies on endophytic bacterial diversity have revealed a large richness of species. Endophytes promote plant growth and yield, suppress pathogens, may help to remove contaminants, solubilize phosphate, or contribute assimilable nitrogen to plants. Some endophytes are seed-borne, but others have mechanisms to colonize the plants that are being studied. Bacterial mutants unable to produce secreted proteins are impaired in the colonization process. Plant genes expressed in the presence of endophytes provide clues as to the effects of endophytes in plants. Molecular analysis showed that plant defense responses limit bacterial populations inside plants. Some human pathogens, such as Salmonella spp., have been found as endophytes, and these bacteria are not removed by disinfection procedures that eliminate superficially occurring bacteria. Delivery of endo-phytes to the environment or agricultural fields should be carefully evaluated to avoid introducing pathogens.
The Tobacco mosaic virus Replicase Protein Disrupts the Localization and Function of Interacting Aux/IAA Proteins
Authors: Padmanabhan Meenu S. , Shiferaw Haiymanot , Culver James N.
Published: May 2007

Previously, we identified a correlation between the interaction of the Tobacco mosaic virus (TMV) 126/183-kDa replicase with the auxin response regulator indole acetic acid (IAA)26/PAP1 and the development of disease symptoms. In this study, the TMV replicase protein is shown to colocalize with IAA26 in the cytoplasm and prevent its accumulation within the nucleus. Furthermore, two additional auxin (Aux)/IAA family members, IAA27 and IAA18, were found to interact with the TMV replicase and displayed alterations in their cellular localization or accumulation that corresponded with their ability to interact with the TMV replicase. In contrast, the localization and accumulation of noninteracting Aux/IAA proteins were unaffected by the presence of the viral replicase. To investigate the effects of the replicase interaction on Aux/IAA function, transgenic plants expressing a proteolysis-resistant IAA26-P108H-green fluorescent protein (GFP) protein were created. Transgenic plants accumulating IAA26-P108H-GFP displayed an abnormal developmental phenotype that included severe stunting and leaf epinasty. However, TMV infection blocked the nuclear localization of IAA26-P108H-GFP and attenuated the developmental phenotype displayed by the transgenic plants. Combined, these findings suggest that TMV-induced disease symptoms can be attributed, in part, to the ability of the viral replicase protein to disrupt the localization and subsequent function of interacting Aux/IAA proteins.
At-4/1, an Interactor of the Tomato spotted wilt virus Movement Protein, Belongs to a New Family of Plant Proteins Capable of Directed Intra- and Intercellular Trafficking
Authors: Paape Martina , Solovyev Andrey G. , Erokhina Tatyana N. , Minina Elena A. , Schepetilnikov Mikhail V. , Lesemann Dietrich-E. , Schiemann Joachim , Morozov Sergey Yu. , Kellmann Jan-W.
Published: May 2007

The Tomato spotted wilt virus (TSWV) encoded NSm movement protein facilitates cell-to-cell spread of the viral genome through structurally modified plasmodesmata. NSm has been utilized as bait in yeast two-hybrid interaction trap screenings. As a result, a protein of unknown function, called At-4/1, was isolated from an Arabidopsis thaliana GAL4 activation domain-tagged cDNA library. Using poly-clonal antibodies against bacterially expressed At-4/1, Western blot analysis of protein extracts isolated from different plant species as well as genome database screenings showed that homologues of At-4/1 seemed to be encoded by many vascular plants. For subcellular localization studies, At-4/1 was fused to green fluorescent protein, and corresponding expression vectors were used in particle bombardment and agroinfiltration assays. Confocal laser scannings revealed that At-4/1 assembled in punctate spots at the cell periphery. The protein accumulated intracellularly in a polarized fashion, appearing in only one-half of a bombarded epidermal cell, and, moreover, moved from cell to cell, forming twin-structured bodies seemingly located at both orifices of the plasmodesmatal pore. In coexpression studies, At-4/1 colocalized with a plant virus movement protein TGBp3 known to reside in endoplasmic reticulum-derived membrane structures located in close vicinity to plasmodesmata. Thus, At-4/1 belongs to a new family of plant proteins capable of directed intra- and intercellular trafficking.
Overexpression of BetS, a Sinorhizobium meliloti High-Affinity Betaine Transporter, in Bacteroids from Medicago sativa Nodules Sustains Nitrogen Fixation During Early Salt Stress Adaptation
Authors: Boscari Alexandre , Van de Sype Ghislaine , Le Rudulier Daniel , Mandon Karine
Published: May 2007

Sinorhizobium meliloti possesses several betaine transporters to cope with salt stress, and BetS represents a crucial high-affinity glycine and proline betaine uptake system involved in the rapid acquisition of betaines by cells subjected to osmotic upshock. Using a transcriptional lacZ (β-galactosidase) fusion, we showed that betS is expressed during the establishment of the symbiosis and in mature nitrogen-fixing nodules. However, neither Nod nor Fix phenotypes were impaired in a betS mutant. BetS is functional in isolated bacteroids, and its activity is strongly activated by high osmolarity. In bacteroids from a betS mutant, glycine betaine and proline betaine uptake was reduced by 85 to 65%, indicating that BetS is a major component of the overall betaine uptake activity in bacteroids in response to osmotic stress. Upon betS overexpression (strain UNA349) in free-living cells, glycine betaine transport was 2.3-fold higher than in the wild-type strain. Interestingly, the accumulation of proline betaine, the endogenous betaine synthesized by alfalfa plants, was 41% higher in UNA349 bacteroids from alfalfa plants subjected to 1 week of salinization (0.3 M NaCl) than in wild-type bacteroids. In parallel, a much better maintenance of nitrogen fixation activity was observed in 7-day-salinized plants nodulated with the over-expressing strain than in wild-type nodulated plants. Taken altogether, these results are consistent with the major role of BetS as an emergency system involved in the rapid uptake of betaines in isolated and in planta osmotically stressed bacteroids of S. meliloti.
Characterization of the Recombinant Forms Arising from aPotato virus XChimeric Virus Infection under RNA Silencing Pressure
Authors: Barajas D. , Tenllado F. , Díaz-Ruíz J. R.
Published: May 2007

Recombination is a frequent phenomenon in RNA viruses whose net result is largely influenced by selective pressures. RNA silencing in plants acts as a defense mechanism against viruses and can be used to engineer virus resistance. Here, we have investigated the influence of RNA silencing as a selective pressure to favor recombinants of PVX-HCT, a chimeric Potato virus X (PVX) vector carrying the helper-component proteinase (HC-Pro) gene from Plum pox virus (PPV). All the plants from two lines expressing a silenced HC-Pro transgene were completely resistant to PPV. However a significant proportion became infected with PVX-HCT. Analysis of viral RNAs accumulating in silenced plants revealed that PVX-HCT escaped silencing-based resistance by removal of the HC-Pro sequences that represented preferential targets for transgene-promoted silencing. The virus vector also tended to lose the HC-Pro insert when infecting transgenic plants containing a nonsilenced HC-Pro transgene or wild-type (wt) Nicotiana benthamiana plants. Nevertheless, loss of HC-Pro sequences was faster in nonsilenced transgenic plants than in wt plants, suggesting the transgene plays a role in promoting a higher selective pressure in favor of recombinant virus versions. These results indicate that the outcome of recombination processes depends on the strength of selection pressures applied to the virus.
Analysis of Nod-Factor-Induced Calcium Signaling in Root Hairs of Symbiotically Defective Mutants of Lotus japonicus
Authors: Miwa Hiroki , Sun Jongho , Oldroyd Giles E. D. , Downie J. Allan
Published: May 2007

Nodulation (Nod)-factor signaling molecules are essential for rhizobia to initiate the nitrogen-fixing symbiotic interaction with legumes. Using a dual dye ratiometric calcium imaging technique, we have shown that 10 nM Nod factor added to roots of Lotus japonicus seedlings induces an intra-cellular calcium increase (calcium flux) that precedes oscillations in intracellular calcium (calcium spiking). The calcium flux was not observed with 1 or 0.1 nM Nod factor, which did induce calcium spiking. The calcium flux was variable in timing of initiation and duration and was observed in approximately half of the root hairs examined. Representatives from 11 complementation groups of symbiotically defective mutants were analyzed for the calcium flux. Mutants from four groups (sym6, ccamk, sym35, and nin) which retained calcium spiking all showed a normal calcium flux. Two classes of mutants (nfr1 and nfr5) lacked both calcium influx and calcium spiking, whereas five classes of mutants (symRK, castor, pollux, nup133, and sym24) defective for calcium spiking retained a calcium flux. There was no correlation between calcium spiking and induction of root hair deformation by Nod factor. We propose that increased bacterial numbers within infection foci in root hairs leads to accumulation of Nod factor to sufficient levels to activate the calcium flux, and this may drive infection thread growth.
Hibiscus chlorotic ringspot virus p27 and Its Isoforms Affect Symptom Expression and Potentiate Virus Movement in Kenaf (Hibiscus cannabinus L.)
Authors: Zhou T. , Fan Z. F. , Li H. F. , Wong S. M.
Published: May 2007

Hibiscus chlorotic ringspot virus (HCRSV), a member of the genus Carmovirus, encodes p27 (27-kDa protein) and two other in-frame isoforms (p25 and p22.5) that are coterminal at the carboxyl end. Only p27, which initiates at the 2570CUG codon, was detected in transfected kenaf (Hibiscus cannabinus L.) protoplasts through fusion to a Flag tag at either its N or C terminus. Subcellular localization of a p27-green fluorescent fusion protein in kenaf epidermal cells showed that it was localized to membrane structures close to cell walls. To study the functions of these proteins, a number of start codon mutants and premature translation termination mutants were constructed. Phenotypic differences were observed between the wild-type virus and these mutants during infection. Infectivity assays on plants indicated that p27 is a determinant of symptom severity. Without p25, appearance of symptoms on systemically infected kenaf leaves was delayed by 4 to 8 days. In a timecourse analysis, Western blot assays revealed that the delay corresponded to retardation in virus systemic movement, which suggested that p25 is probably involved in virus systemic movement. Mutations disrupting expression of p22.5 did not affect symptoms or virus movement.
Genome-Wide Gene Expression Analysis of Pseudomonas syringae pv. tomato DC3000 Reveals Overlapping and Distinct Pathways Regulated by hrpL and hrpRS
Authors: Lan Lefu , Deng Xin , Zhou Jianmin , Tang Xiaoyan
Published: May 2007

Pseudomonas syringae pv. tomato DC3000 is a model pathogen infecting tomato and Arabidopsis plants. Genes encoding the type III secretion system and substrate proteins (collectively called TTSS genes) of this bacterium are induced in plants and in minimal medium (MM). The induction of TTSS genes is mediated by HrpL, an alternative sigma factor recognizing the hrp box in the promoter of TTSS genes. The transcription of hrpL is activated by HrpR and HrpS, two homologous DNA-binding proteins encoded by the hrpRS operon. Microarray analysis was conducted to evaluate the DC3000 genes regulated by hrpL and hrpRS in MM. The analysis identified a number of novel hrpL-activated genes with a putative TTSS-independent function. Genes regulated by hrpL were mostly regulated by hrpRS in the same manner, but a large number of genes regulated by hrpRS were hrpL-independent, indicating that hrpL represents one branch of the regulatory pathways downstream of hrpRS. The induction of the TTSS genes was associated with downregulation of the housekeeping genes, indicating that the activation of the TTSS has a cost on the basic cellular activities. The novel genes and pathways identified by the microarray provide new insight into the bacterial functions coordinating with the TTSS.
Metabolite Profiles of Nodulated Alfalfa Plants Indicate That Distinct Stages of Nodule Organogenesis Are Accompanied by Global Physiological Adaptations
Authors: Barsch Aiko , Tellström Verena , Patschkowski Thomas , Küster Helge , Niehaus Karsten
Published: May 2007

An effective symbiosis between Sinorhizobium meliloti and its host plant Medicago sativa is dependent on a balanced physiological interaction enabling the microsymbiont to fix atmospheric nitrogen. Maintenance of the symbiotic interaction is regulated by still poorly understood control mechanisms. A first step toward a better understanding of nodule metabolism was the determination of characteristic metabolites for alfalfa root nodules. Furthermore, nodules arrested at different developmental stages were analyzed in order to address metabolic changes induced during the progression of nodule formation. Metabolite profiles of bacteroid-free pseudonodule extracts indicated that early nodule developmental processes are accompanied by photosynthate translocation but no massive organic acid formation. To determine metabolic adaptations induced by the presence of nonfixing bacteroids, nodules induced by mutant S. meliloti strains lacking the nitrogenase protein were analyzed. The bacteroids are unable to provide ammonium to the host plant, which is metabolically reflected by reduced levels of characteristic amino acids involved in ammonium fixation. Elevated levels of starch and sugars in Fix¯ nodules provide strong evidence that plant sanctions preventing a transformation from a symbiotic to a potentially parasitic interaction are not strictly realized via photo-synthate supply. Instead, metabolic and gene expression data indicate that alfalfa plants react to nitrogen-fixation-deficient bacteroids with a decreased organic acid synthesis and an early induction of senescence. Noneffective symbiotic interactions resulting from plants nodulated by mutant rhizobia also are reflected in characteristic metabolic changes in leaves. These are typical for nitrogen deficiency, but also highlight metabolites potentially involved in sensing the N status.
Global Impact: Elucidating Plant Responses to Viral Infection
Authors: Whitham Steven A. , Yang Chunling , Goodin Michael M.
Published: May 2007

Viruses induce a variety of responses in host cells that are mediated by perturbation of different signaling pathways. Advances in our understanding of the functions of viral proteins, plant biology in general, as well as technologies for profiling gene expression have converged in recent years to provide new insight into the events occurring inside susceptible and resistant host cells in response to virus infection. These effects range from nonspecific changes in gene expression due to the general accumulation of viral proteins to those responses that are initiated by the specific interactions between virus and host proteins. Here, we discuss a variety of expression profiling methods and approaches that have been used to study the effects of viruses on host transcriptomes. These studies have identified distinct sets of genes that have altered expression profiles in response to viruses, including stress- and defense-related genes. The activities of viral RNA silencing suppressors and interference with hormone signaling or biogenesis also influence plant gene expression and lead to developmental abnormalities.
Innate Nonhost Immunity in Barley to Different Heterologous Rust Fungi Is Controlled by Sets of Resistance Genes with Different and Overlapping Specificities
Authors: Jafary Hossein , Szabo Les J. , Niks Rients E.
Published: May 2007

We developed an evolutionary relevant model system, barley-Puccini rust fungi, to study the inheritance and specificity of plant factors that determine to what extent innate nonhost immunity can be suppressed. A mapping population was developed from a cross between an experimental barley line (Suspired) with exceptional susceptibility to several heterogonous (nonhost) rust fungi and regular, immune, cv. Veda. Seedlings were inoculated with five heterogonous and two homologous (host) species of rust fungi. Resistance segregated quantitatively for each of the rust fungi. In total, 18 chromosomal regions were implicated. For each rust species, a different set of genes was effective. Of the 18 chromosomal regions, 11 were significantly effective to only one rust species and 7 were effective to more than one rust species, implying genetic linkage or pleiotropy. One resistance (R) gene for hypersensitive resistance to Puccinia hordei-secalini was mapped, suggesting occasional contribution of R genes to nonhost resistance in barley. Quantitative trait loci (QTLs) with effects to multiple rust fungi did not tend to be particularly effective to rust species that were phylogenetically related, as determined from their internal transcribed spacer sequence. We suggest that the QTLs described here play a role as specific and quantitative recognition factors that are specifically negated by the rust to successfully suppress innate immunity.
Phytophthora Genomics: The Plant Destroyers' Genome Decoded
Authors: Govers Francine , Gijzen Mark
Published: May 2007

The year 2004 was an exciting one for the Phytophthora research community. The United States Department of Energy Joint Genome Institute (JGI) completed the draft genome sequence of two Phytophthora species, Phytophthora sojae and Phytophthora ramorum. In August of that year over 50 people gathered at JGI in Walnut Creek, California, for an annotation jamboree and searched for the secrets and surprises that the two genomes have in petto. This culminated in a paper in Science in September of this year describing the highlights of the sequencing project and emphasizing the power of having the genome sequences of two closely related organisms. This MPMI Focus issue on Phytophthora genomics contains a number of more specialized manuscripts centered on gene annotation and genome organization, and complemented with manuscripts that rely on genomics resources.
Cladosporium fulvum Avr4 Protects Fungal Cell Walls Against Hydrolysis by Plant Chitinases Accumulating During Infection
Authors: van den Burg Harrold A. , Harrison Stuart J. , Joosten Matthieu H. A. J. , Vervoort Jacques , de Wit Pierre J. G. M.
Published: May 2007

Resistance against the leaf mold fungus Cladosporium fulvum is mediated by the tomato Cf proteins which belong to the class of receptor-like proteins and indirectly recognize extracellular avirulence proteins (Avrs) of the fungus. Apart from triggering disease resistance, Avrs are believed to play a role in pathogenicity or virulence of C. fulvum. Here, we report on the avirulence protein Avr4, which is a chitin-binding lectin containing an invertebrate chitin-binding domain (CBM14). This domain is found in many eukaryotes, but has not yet been described in fungal or plant genomes. We found that interaction of Avr4 with chitin is specific, because it does not interact with other cell wall polysaccharides. Avr4 binds to chitin oligomers with a minimal length of three N-acetyl glucosamine residues. In vitro, Avr4 protects chitin against hydrolysis by plant chitinases. Avr4 also binds to chitin in cell walls of the fungi Trichoderma viride and Fusarium solani f. sp. phaseoli and protects these fungi against normally deleterious concentrations of plant chitinases. In situ fluorescence studies showed that Avr4 also binds to cell walls of C. fulvum during infection of tomato, where it most likely protects the fungus against tomato chitinases, suggesting that Avr4 is a counter-defensive virulence factor.
Identification of Symbiotically Defective Mutants of Lotus japonicus Affected in Infection Thread Growth
Authors: Lombardo Fabien , Heckmann Anne B. , Miwa Hiroki , Perry Jillian A. , Yano Koji , Hayashi Makoto , Parniske Martin , Wang Trevor L. , Downie J. Allan
Published: May 2007

During the symbiotic interaction between legumes and rhizobia, the host cell plasma membrane and associated plant cell wall invaginate to form a tunnel-like infection thread, a structure in which bacteria divide to reach the plant root cortex. We isolated four Lotus japonicus mutants that make infection pockets in root hairs but form very few infection threads after inoculation with Mesorhizobium loti. The few infection threads that did initiate in the mutants usually did not progress further than the root hair cell. These infection-thread deficient (itd) mutants were unaffected for early symbiotic responses such as calcium spiking, root hair deformation, and curling, as well as for the induction of cortical cell division and the arbuscular mycorrhizal symbiosis. Complementation tests and genetic mapping indicate that itd2 is allelic to Ljsym7, whereas the itd1, itd3, and itd4 mutations identified novel loci. Bacterial release into host cells did occur occasionally in the itd1, itd2, and itd3 mutants suggesting that some infections may succeed after a long period and that infection of nodule cells could occur normally if the few abnormal infection threads that were formed reached the appropriate nodule cells.
Viroid: A Useful Model for Studying the Basic Principles of Infection and RNA Biology
Authors: Ding Biao , Itaya Asuka
Published: May 2007

Viroids are small, circular, noncoding RNAs that currently are known to infect only plants. They also are the smallest self-replicating genetic units known. Without encoding proteins and requirement for helper viruses, these small RNAs contain all the information necessary to mediate intracellular trafficking and localization, replication, systemic trafficking, and pathogenicity. All or most of these functions likely result from direct interactions between distinct viroid RNA structural motifs and their cognate cellular factors. In this review, we discuss current knowledge of these RNA motifs and cellular factors. An emerging theme is that the structural simplicity, functional versatility, and experimental tractability of viroid RNAs make viroid-host interactions an excellent model to investigate the basic principles of infection and further the general mechanisms of RNA-templated replication, intracellular and intercellular RNA trafficking, and RNA-based regulation of gene expression. We anticipate that significant advances in understanding viroid-host interactions will be achieved through multifaceted secondary and tertiary RNA structural analyses in conjunction with genetic, biochemical, cellular, and molecular tools to characterize the RNA motifs and cellular factors associated with the processes leading to systemic infection.
Identification and Characterization of Plant Genes Involved inAgrobacterium-Mediated Plant Transformation by Virus-Induced Gene Silencing
Authors: Anand Ajith , Vaghchhipawala Zarir , Ryu Choong-Min , Kang Li , Wang Keri , del-Pozo Olga , Martin Gregory B. , Mysore Kirankumar S.
Published: May 2007

Genetic transformation of plant cells by Agrobacterium tu-mefaciens represents a unique case of trans-kingdom sex requiring the involvement of both bacterial virulence proteins and plant-encoded proteins. We have developed in planta and leaf-disk assays in Nicotiana benthamiana for identifying plant genes involved in Agrobacterium-mediated plant transformation using virus-induced gene silencing (VIGS) as a genomics tool. VIGS was used to validate the role of several genes that are either known or speculated to be involved in Agrobacterium-mediated plant transformation. We showed the involvement of a nodulin-like protein and an alpha-expansin protein (α-Exp) during Agrobacterium infection. Our data suggest that α-Exp is involved during early events of Agrobacterium-mediated transformation but not required for attaching A. tumefaciens. By employing the combination of the VIGS-mediated forward genetics approach and an in planta tumorigenesis assay, we identified 21 ACG (altered crown gall) genes that, when silenced, produced altered crown gall phenotypes upon infection with a tumorigenic strain of A. tumefaciens. One of the plant genes identified from the screening, Histone H3 (H3), was further characterized for its biological role in Agrobacterium-mediated plant transformation. We provide evidence for the role of H3 in transfer DNA integration. The data presented here suggest that the VIGS-based approach to identify and characterize plant genes involved in genetic transformation of plant cells by A. tumefaciens is simple, rapid, and robust and complements other currently used approaches.
Involvement of the Octadecanoid Pathway in Bluegreen Aphid Resistance in Medicago truncatula
Authors: Gao Ling-Ling , Anderson Jonathan P. , Klingler John P. , Nair Ramakrishnan M. , Edwards Owain R. , Singh Karam B.
Published: May 2007

Aphids are major insect pests of plants that feed directly from the phloem. We used the model legume Medicago truncatula Gaert. (barrel medic) to elucidate host resistance to aphids and identified a single dominant gene which confers resistance to Acyrthosiphon kondoi Shinji (bluegreen aphid). To understand how this gene conditions resistance to bluegreen aphid, transcription profiling of 23 defense-related genes representing various signaling pathways was undertaken using a pair of near-isogenic lines that are susceptible or resistant to bluegreen aphid. All salicylic acid- and ethylene-responsive genes tested were induced by blue-green aphid in resistant and susceptible plants, although there were some differences in the magnitude and kinetics of the induction. In contrast, 10 of 13 genes associated with the octadecanoid pathway were induced exclusively in the resistant plants following bluegreen aphid infestation. These results are in contrast to plant-pathogen interactions where similar sets of defense genes typically are induced in compatible interactions, but to a lesser degree and later than in incompatible interactions. Treatment of susceptible plants with methyl jasmonate reduced bluegreen aphid infestation but not to the same levels as the resistant line. Together, these results strongly suggest that the octa-decanoid pathway is important for this naturally derived aphid resistance trait.
The HrpNea Harpin from Erwinia amylovora Triggers Differential Responses on the Nonhost Arabidopsis thaliana Cells and on the Host Apple Cells
Authors: Reboutier David , Frankart Cécile , Briand Joël , Biligui Bernadette , Laroche Sandrine , Rona Jean-Pierre , Barny Marie-Anne , Bouteau François
Published: May 2007

Erwinia amylovora is a gram-negative necrogenic bacterium causing fire blight of the Maloideae subfamily of Rosaceae such as apple and pear. It provokes progressive necrosis in aerial parts of susceptible host plants (compatible interaction) and a hypersensitive reaction (HR) when infiltrated in nonhost plants (incompatible interaction). The HrpNea harpin is a type three secretion system effector secreted by E. amylovora. This protein is involved in pathogenicity and HR-eliciting capacity of E. amylovora. In the present study, we showed that, in nonhost Arabidopsis thaliana cells, purified HrpNea induces cell death and H2O2 production, two nonhost resistance responses, but failed to induce such responses in host MM106 apple cells. Moreover, HrpNea induced an increase in anion current in host MM106 apple cells, at the opposite of the decrease of anion current previously shown to be necessary to induce cell death in nonhost A. thaliana cells. These results suggest that HrpNea induced different signaling pathways, which could account for early induced compatible or incompatible interaction development.
Transcriptional Analysis of Complementary Sense Genes in Spinach curly top virus and Functional Role of C2 in Pathogenesis
Authors: Baliji Surendranath , Sunter Janet , Sunter Garry
Published: May 2007

Spinach curly top virus (SCTV), the fifth characterized Curtovirus species belonging to the family Geminiviridae, is an agriculturally significant plant pathogen representing an emerging disease threat in the southern United States. The SCTV genome comprises a single DNA chromosome of approximately 3.0 kb, with the potential to code for seven proteins larger than 10 kDa but which relies extensively on the host for replication and transcription of its genome. In this study, we have identified viral and complementary sense transcripts in SCTV-infected plants, confirming a bidirectional transcription strategy for SCTV. The most abundant RNA maps to the virion sense (1.1-kb transcript) and is comparable in size and location to that observed in Beet curly top virus (BCTV). Two complementary sense transcripts (1.7 and 0.7 kb) were identified in SCTV-infected plants. The large, 1.7-kb transcript is comparable in size and position to that identified in BCTV and several begomoviruses and most likely encodes the C1 protein. Both complementary sense RNAs could potentially direct expression of C2 and C3 from polycistronic mRNAs. A mutation in the C2 gene of SCTV results in expression of a truncated protein of 38 amino acids that is capable of interacting with two cellular kinases, AKIN11 and ADK2, and the resulting mutant virus remains highly infectious. A second mutant virus can only express the first three amino acids of the C2 protein and is unable to interact with the same kinases. However, this mutant virus still remains infectious, although a reduction in infectivity and symptom severity was seen in both Arabidopsis and spinach. A possible relationship between the interaction of C2 with AKIN11 and ADK2 and disease severity is presented.
Bacillus megaterium Rhizobacteria Promote Growth and Alter Root-System Architecture Through an Auxin- and Ethylene-Independent Signaling Mechanism in Arabidopsis thaliana
Authors: López-Bucio José , Campos-Cuevas Juan Carlos , Hernández-Calderón Erasto , Velásquez-Becerra Crisanto , Farías-Rodríguez Rodolfo , Macías-Rodríguez Lourdes Iveth , Valencia-Cantero Eduardo
Published: May 2007

Soil microorganisms are critical players in plant-soil interactions at the rhizosphere. We have identified a Bacillus megaterium strain that promoted growth and development of bean (Phaseolus vulgaris) and Arabidopsis thaliana plants. We used Arabidopsis thaliana as a model to characterize the effects of inoculation with B. megaterium on plant-growth promotion and postembryonic root development. B. megaterium inoculation caused an inhibition in primary-root growth followed by an increase in lateral-root number, lateral-root growth, and root-hair length. Detailed cellular analyses revealed that primary root-growth inhibition was caused both by a reduction in cell elongation and by reduction of cell proliferation in the root meristem. To study the contribution of auxin and ethylene signaling pathways in the alterations in root-system architecture elicited by B. megaterium, a suite of plant hormone mutants of Arabidopsis, including aux1-7, axr4-1, eir1, etr1, ein2, and rhd6, defective in either auxin or ethylene signaling, were evaluated for their responses to inoculation with this bacteria. When inoculated, all mutant lines tested showed increased biomass production. Moreover, aux1-7 and eir1, which sustain limited root-hair and lateral-root formation when grown in uninoculated medium, were found to increase the number of lateral roots and to develop long root hairs when inoculated with B. megaterium. The ethylene-signaling mutants etr1 and ein2 showed an induction in lateral-root formation and root-hair growth in response to bacterial inoculation. Taken together, our results suggest that plant-growth promotion and root-architectural alterations by B. megaterium may involve auxin- and-ethylene independent mechanisms.
Spatial Analysis ofArabidopsis thalianaGene Expression in Response toTurnip mosaic virusInfection
Authors: Yang Chunling , Guo Rong , Jie Fei , Nettleton Dan , Peng Jiqing , Carr Tyrell , Yeakley Joanne M. , Fan Jian-Bing , Whitham Steven A.
Published: May 2007

Virus-infected leaf tissues comprise a heterogeneous mixture of cells at different stages of infection. The spatial and temporal relationships between sites of virus accumulation and the accompanying host responses, such as altered host gene expression, are not well defined. To address this issue, we utilized Turnip mosaic virus (TuMV) tagged with the green fluorescent protein to guide the dissection of infection foci into four distinct zones. The abundance of Arabidopsis thaliana mRNA transcripts in each of the four zones then was assayed using the Arabidopsis ATH1 GeneChip oligonucleotide microarray (Affymetrix). mRNA transcripts with significantly altered expression profiles were determined across gradients of virus accumulation spanning groups of cells in and around foci at different stages of infection. The extent to which TuMV-responsive genes were up- or downregulated primarily correlated with the amount of virus accumulation regardless of gene function. The spatial analysis also allowed new suites of coordinately regulated genes to be identified that are associated with chloroplast functions (decreased), sulfate assimilation (decreased), cell wall extensibility (decreased), and protein synthesis and turnover (induced). The functions of these downregulated genes are consistent with viral symptoms, such as chlorosis and stunted growth, providing new insight into mechanisms of pathogenesis.
Molecular and Pathotypic Characterization of New Xanthomonas oryzae Strains from West Africa
Authors: Gonzalez Carolina , Szurek Boris , Manceau Charles , Mathieu Thierry , Séré Yacouba , Verdier Valérie
Published: May 2007

DNA polymorphism analysis and pathogenicity assays were used to characterize strains of Xanthomonas oryzae pv. oryzae and Xanthomonas oryzae pv. oryzicola collected from rice leaves in West Africa. Restriction fragment length polymorphism (RFLP), repetitive sequence-based polymerase chain reaction, fluorescent amplified fragment-length polymorphism (FAFLP) analyses were assessed for molecular characterization, while pathogenicity was tested by leaf clipping and leaf infiltration. Dendrograms were generated for the data sets obtained from RFLP analysis and repetitive polymerase chain reaction suggesting that the interrelationships between strains were dependent on the technique used. In all cases, data showed that African strains of X. oryzae pv. oryzae form a group genetically distant from Asian strains. FAFLP analyses separated the X. oryzae strains into three groups with significant bootstrap values. A specific and intriguing feature of African strains of X. oryzae pv. oryzae is a reduction in the number of insertion sequence elements and transcription activator-like (avrBs3/pthA) effector genes, based on the molecular markers employed in the study. In addition, pathogenicity assays conducted with African strains of X. oryzae pv. oryzae on a series of nearly isogenic lines (NILs) identified three new races. Finally, leaf infiltration assays revealed the capacity of African strains of X. oryzae pv. oryzae to induce a nonhost hypersensitive response in Nicotiana benthamiana, in contrast with Asian X. oryzae pv. oryzae and X. oryzae pv. oryzicola strains. Our results reveal substantial differences between genomic characteristics of Asian and African strains of X. oryzae pv. oryzae.
Refinement of the Xanthomonas campestris pv. vesicatoria hrpD and hrpE Operon Structure
Authors: Weber Ernst , Berger Carolin , Bonas Ulla , Koebnik Ralf
Published: May 2007

The plant-pathogenic bacterium Xanthomonas campestris pv. vesicatoria possesses a type III secretion (T3S) system which is encoded in the 23-kb hypersensitive response and pathogenicity (hrp) gene cluster. The T3S system is essential for pathogenicity in susceptible hosts and the induction of the hypersensitive response in resistant plants. In this study, we revisited the operon structure of the right part of the hrp gene cluster. Based on complementation experiments of transposon insertions and reverse-transcription polymerase chain reaction analyses, the hrpD operon contains hrcQ, hrcR, hrcS, and hpaA, whereas hrcD, hrpD6, and hrpE belong to the hrpE operon. We determined the transcriptional start site of the hrpE operon and showed that there is a promoter upstream of hrcD containing a plant-inducible promoter box. Conserved secondary mRNA structures in the intergenic region between hrpD6 and hrpE suggest a posttranscriptional regulated expression of hrpE. Based on comparisons of different hrp gene clusters and the analysis of evolutionary rates, we propose that the hrpE transcriptional unit was integrated into the hrp gene cluster at a later time.
Description of the Colonization of a Gnotobiotic Tomato Rhizosphere by Pseudomonas fluorescens Biocontrol Strain WCS365, Using Scanning Electron Microscopy
Authors: Chin-A-Woeng Thomas F. C. , de Priester Wessel , van der Bij Arjan J. , Lugtenberg Ben J. J.
Published: May 2007

To study colonization of the tomato root system, we previously have described a gnotobiotic quartz sand system, in which seedlings inoculated with one or two bacterial strains were allowed to grow. Here we present a scanning electron microscope description of the colonization of the tomato root system by Pseudomonas fluorescens biocontrol strain WCS365, with emphasis on spatial-temporal colonization patterns, based on an improved scanning electron microscopy procedure. Upon inoculation of the germinated seed, proliferation on the seed coat was observed for 2 to 3 days. Within 1 to 3 days, micro-colonies developed, mainly at the root base. Most micro-colonies were localized in junctions between epidermal root cells, whereas others were found in indented parts of the epidermal surface. Downward to the root tip, only single bacterial cells were found. Colonization progressed down the root, initially as single cells. A semi-transparent film appeared to enclose the root surface and micro-colonies present on the root. After 7 days, micro-colonies had developed at positions where only single cells were observed previously and distribution of the bacteria along the root varied from ≈106 CFU per cm of root near the root base to ≈102 to 103 CFU per cm of root near the root tip. Similar colonization patterns were found for the P. fluorescens biocontrol strains CHA0 and F113, and P. putida strain WCS358, as well as for four species that have repeatedly been isolated from tomato roots from a commercial tomato field near Granada, Spain. In contrast, four Rhizobium strains and one Acinetobacter radioresistens strain showed poor colonization and micro-colonies were not observed. Based on the described data, we present a model for colonization of the deeper root parts after seed inoculation by P. fluorescens biocontrol strains, in which single cells occasionally establish on a deeper root section where they sometimes develop into micro-colonies. We hypothesize that micro-colonies are the sites where the intracellular N-acyl-L-homoserine lactone concentration is sufficiently high to cause maximal production of biocontrol factors such as antibiotics and exoenzymes and that micro-colonies explain the relatively high conjugation frequency observed between pseudomonads in the rhizosphere.
rosR, a Determinant of Nodulation Competitiveness in Rhizobium etli
Authors: Bittinger Mark A. , Milner Jocelyn L. , Saville Barry J. , Handelsman Jo
Published: May 2007

We previously described a Tn5 mutant of Rhizobium etli strain CE3, designated CE3003, that is decreased in nodulation competitiveness, reduced in competitive growth in the rhizosphere, and has a hydrophobic cell surface (R. S. Araujo, E. A. Robleto, and J. Handelsman, Appl. Environ. Microbiol., 60:1430–1436, 1994). To determine the molecular basis for the mutant phenotypes, we identified a 1.2-kb fragment of DNA derived from the parent that restored the wild-type phenotypes to the mutant. DNA sequence analysis indicated that this 1.2-kb fragment contained a single open reading frame that we designated rosR. The Tn5 insertion in CE3003 was within rosR. We constructed a derivative of CE3 that contained a deletion in rosR, and this mutant was phenotypically indistinguishable from CE3003 in cell surface and competitive characteristics. Based on the nucleotide sequence, the deduced RosR amino acid sequence is 80% identical to that of the Ros protein from Agrobacterium tumefaciens and the MucR protein from Rhizobium meliloti. Both Ros and MucR are transcriptional repressors that contain a putative zinc-finger DNA-binding domain. This study defines a gene, rosR, that is homologous to a family of transcriptional regulators and contributes to nodulation competitiveness of R. etli. Moreover, we established that a single gene affects nodulation competitiveness, competitive growth in the rhizosphere, and cell surface hydrophobicity.
Trifolitoxin Production in Rhizobium etli Strain CE3 Increases Competitiveness for Rhizosphere Colonization and Root Nodulation of Phaseolus vulgaris in Soil
Authors: Robleto Eduardo A. , Scupham Alexandra J. , Triplett Eric W.
Published: May 2007

The effect of trifolitoxin (TFX) production by Rhizobium etli on rhizosphere colonization and competition for nodulation in soil conditions was determined. TFX is a potent peptide antibiotic made by Rhizobium leguminosarum bv. trifolii T24 that inhibits many α-proteobacteria (E. W. Triplett, B. T. Breil, and G. A. Splitter, Appl. Environ. Microbiol. 60:4163–4166, 1994). Seeds of Phaseolus vulgaris were inoculated with a TFX-sensitive reference strain and either of two isogenic strains that differ only in their ability to produce TFX. The pair of strains were inoculated at different ratios in sterile and nonsterile soil. The representation of the strains in the rhizosphere and nodules was determined at 96 h after inoculation and 3 weeks after planting, respectively. The TFX-producing strain was significantly more competitive for both phenotypes versus the TFX-sensitive strain, compared with the TFX-nonproducing strain versus the TFX-sensitive strain. These results show that nodule occupancy by inoculant strains, often displaced from the nodules by indigenous strains, can be increased by addition of the TFX production phenotype to R. etli in plants grown in either sterile or nonsterile soil. Also, this work shows the efficacy of the TFX system for the first time on a legume host with determinant nodules.
A Hormogonium Regulating Locus,hrmUA, of the CyanobacteriumNostoc punctiformeStrain ATCC 29133 and its Response to an Extract of a Symbiotic Plant PartnerAnthoceros punctatus
Authors: Cohen Michael F. , Meeks John C.
Published: May 2007

Transposon-generated mutant strain UCD 328 of Nostoc punctiforme strain ATCC 29133 has a phenotype of an increased sensitivity to a hormogonium-inducing factor exuded by a symbiotic plant partner, Anthoceros punctatus, and an initial increased hormogonium-dependent infection of the plant. Sequence analysis showed that the transposition site in strain UCD 328 lies within a 1,251-bp open reading frame (ORF), designated hrmA, that displays no significant similarity to known database sequences. A second, 837-bp ORF (hrmU) ends 2 bp 5′ from the start of hrmA and has the signature sequences belonging to a family of NAD(P)H-dependent oxidoreductases. Strains having insertional mutations in hrmU or hrmA reproduce the strain UCD 328 phenotype. Transcriptional fusions of luxAB to hrmU or hrmA show an 8- to 10-fold peak increase in luciferase activity 13 to 20 h after the start of incubation in the presence of an aqueous extract of A. punctatus. A promoter induced by the extract was deduced to be between 2.0 to 3.4 kb from the translational start of hrmU. A multicopy plasmid that contains hrmUA within a 6.2-kb fragment conferred an increased infection phenotype on wild-type N. punctiforme 29133. This plasmid and another plasmid containing 4.4 kb of DNA 5′ of the transposition site prevented extract-dependent induction of hrmA-luxAB transcription in strain UCD 328, implicating titration of some trans-activator(s) by the cloned fragments. We hypothesize a role for hrmUA in the inhibition of hormogonium formation by the metabolism of an unknown hormogonium-regulating metabolite.
Effect of Disruption of a Cutinase Gene (cutA) on Virulence and Tissue Specificity of Fusarium solani f. sp. cucurbitae race 2 Toward Cucurbita maxima and C. moschata
Authors: Crowhurst Ross N. , Binnie Stephanie J. , Bowen Joanna K. , Hawthorne Brian T. , Plummer Kim M. , Rees-George Jonathan , Rikkerink Erik H. A. , Templeton Matthew D.
Published: May 2007

A 3.9-kb genomic DNA fragment from the cucurbit pathogen Fusarium solani f. sp. cucurbitae race 2 was cloned. Sequence analysis revealed an open reading frame of 690 nucleotides interrupted by a single 51-bp intron. The nucleotide and predicted amino acid sequences showed 92 and 98% identity, respectively, to those of the cutA gene of the pea pathogen F. solani f. sp. pisi. A gene replacement vector was constructed and used to generate cutA - mutants that were detected with a polymerase chain reaction (PCR) assay. Seventy-one cutA- mutants were identified among the 416 transformants screened. Vector integration was assessed by Southern analysis in 23 of these mutants. PCR and Southern analysis data showed the level of homologous integration was 14%. Disruption of the cutA locus in mutants was confirmed by RNA gel blot hybridization. Neither virulence on Cucurbita maxima cv. Delica at any of six different inoculum concentrations, nor pathogenicity on intact fruit of four different species or cultivars of cucurbit or hypocotyl tissue of C. maxima cv. Crown, was found to be affected by disruption of the cutA gene.