bluuebunny/crossref_metadata_2025 huggingface dataset:
The ntrPR Operon of Sinorhizobium meliloti Is Organized and Functions as a Toxin-Antitoxin Module
Authors: Bodogai Monica , Ferenczi Szilamér , Bashtovyy Denys , Miclea Paul , Papp Péter , Dusha Ilona
Published: May 2007

The chromosomal ntrPR operon of Sinorhizobium meliloti encodes a protein pair that forms a toxin-antitoxin (TA) module, the first characterized functional TA system in Rhizobiaceae. Similarly to other bacterial TA systems, the toxin gene ntrR is preceded by and partially overlaps with the antitoxin gene ntrP. Based on protein homologies, the ntrPR operon belongs to the vapBC family of TA systems. The operon is negatively autoregulated by the NtrPNtrR complex. Promoter binding by NtrP is weak; stable complex formation also requires the presence of NtrR. The N-terminal part of NtrP is responsible for the interaction with promoter DNA, whereas the C-terminal part is required for protein-protein interactions. In the promoter region, a direct repeat sequence was identified as the binding site of the NtrPNtrR complex. NtrR expression resulted in the inhibition of cell growth and colony formation; this effect was counteracted by the presence of the antitoxin NtrP. These results and our earlier observations demonstrating a less effective downregulation of a wide range of symbiotic and metabolic functions in the ntrR mutant under micro-oxic conditions and an increased symbiotic efficiency with the host plant alfalfa suggest that the ntrPR module contributes to adjusting metabolic levels under symbiosis and other stressful conditions.
Sm1, a Proteinaceous Elicitor Secreted by the Biocontrol Fungus Trichoderma virens Induces Plant Defense Responses and Systemic Resistance
Authors: Djonović Slavica , Pozo Maria J. , Dangott Lawrence J. , Howell Charles R. , Kenerley Charles M.
Published: May 2007

The soilborne filamentous fungus Trichoderma virens is a biocontrol agent with a well-known ability to produce antibiotics, parasitize pathogenic fungi, and induce systemic resistance in plants. Even though a plant-mediated response has been confirmed as a component of bioprotection by Trichoderma spp., the molecular mechanisms involved remain largely unknown. Here, we report the identification, purification, and characterization of an elicitor secreted by T. virens, a small protein designated Sm1 (small protein 1). Sm1 lacks toxic activity against plants and microbes. Instead, native, purified Sm1 triggers production of reactive oxygen species in monocot and dicot seedlings, rice, and cotton, and induces the expression of defense-related genes both locally and systemically in cotton. Gene expression analysis revealed that SM1 is expressed throughout fungal development under different nutrient conditions and in the presence of a host plant. Using an axenic hydroponic system, we show that SM1 expression and secretion of the protein is significantly higher in the presence of the plant. Pretreatment of cotton cotyledons with Sm1 provided high levels of protection to the foliar pathogen Colletotrichum sp. These results indicate that Sm1 is involved in the induction of resistance by Trichoderma spp. through the activation of plant defense mechanisms.
Synergistic Interactions of the Plant Cell Death Pathways Induced by Phytophthora infestans Nep1-Like Protein PiNPP1.1 and INF1 Elicitin
Authors: Kanneganti Thirumala-Devi , Huitema Edgar , Cakir Cahid , Kamoun Sophien
Published: May 2007

Cell death plays a ubiquitous role in plant-microbe interactions, given that it is associated with both susceptible and resistance interactions. A class of cell death-inducing proteins, termed Nep1-like proteins (NLPs), has been reported in bacteria, fungi, and oomycetes. These proteins induce nonspecific necrosis in a variety of dicotyledonous plants. Here, we describe three members of the NLP family from the oomycete Phytophthora infestans (PiNPP1.1, PiNPP1.2, and PiNPP1.3). Using agroinfection with a binary Potato virus X vector, we showed that PiNPP1.1 induces cell death in Nicotiana benthamiana and the host plant tomato. Expression analyses indicated that PiNPP1.1 is up-regulated during late stages of infection of tomato by P. infestans. We compared PiNPP1.1 necrosis-inducing activity to INF1 elicitin, a well-studied protein that triggers the hypersensitive response in Nicotiana spp. Using virus-induced gene silencing, we showed that the cell death induced by PiNPP1.1 is dependent on the ubiquitin ligase-associated protein SGT1 and the heat-shock protein HSP90. In addition, cell death triggered by PiNPP1.1 but not that by INF1 was dependent on the defense-signaling proteins COI1, MEK2, NPR1, and TGA2.2, suggesting distinct signaling requirements. Combined expression of PiNPP1.1 and INF1 in N. benthamiana resulted in enhanced cell death, suggesting synergistic interplay between the two cell-death responses. Altogether, these results point to potentially distinct but interacting cell-death pathways induced by PiNPP1.1 and INF1 in plants.
GacS-Dependent Production of 2R, 3R-Butanediol by Pseudomonas chlororaphis O6 Is a Major Determinant for Eliciting Systemic Resistance Against Erwinia carotovora but not Against Pseudomonas syringae pv. tabaci in Tobacco
Authors: Han Song Hee , Lee Seung Je , Moon Jae Hak , Park Keun Hyung , Yang Kwang Yeol , Cho Baik Ho , Kim Kil Yong , Kim Yong Whan , Lee Myung Chul , Anderson Anne J. , Kim Young Cheol
Published: May 2007

Root colonization by a plant-beneficial rhizobacterium, Pseudomonas chlororaphis O6, induces disease resistance in tobacco against leaf pathogens Erwinia carotovora subsp. carotovora SCC1, causing soft-rot, and Pseudomonas syringae pv. tabaci, causing wildfire. In order to identify the bacterial determinants involved in induced systemic resistance against plant diseases, extracellular components produced by the bacterium were fractionated and purified. Factors in the culture filtrate inducing systemic resistance were retained in the aqueous fraction rather than being partitioned into ethyl acetate. Fractionation on high-performance liquid chromatography followed by nuclear magnetic resonance mass spectrometry analysis identified the active compound as 2R, 3R-butanediol. 2R, 3R butanediol induced systemic resistance in tobacco to E. carotovora subsp. carotovora SCC1, but not to P. syringae pv. tabaci. Treatment of tobacco with the volatile 2R, 3R-butanediol enhanced aerial growth, a phenomenon also seen in plants colonized by P. chlororaphis O6. The isomeric form of the butanediol was important because 2S, 3S-butandiol did not affect the plant. The global sensor kinase, GacS, of P. chlororaphis O6 was a key regulator for induced systemic resistance against E. carotovora through regulation of 2R, 3R-butanediol production. This is the first report of the production of these assumed fermentation products by a pseudomonad and the role of the sensor kinase GacS in production of 2R, 3R-butanediol.
Mutations in DMI3 and SUNN Modify the Appressorium-Responsive Root Proteome in Arbuscular Mycorrhiza
Authors: Amiour Nardjis , Recorbet Ghislaine , Robert Franck , Gianinazzi Silvio , Dumas-Gaudot Eliane
Published: May 2007

Modification of the Medicago truncatula root proteome during the early stage of arbuscular mycorrhizal symbiosis was investigated by comparing, using two-dimensional electrophoresis, the protein patterns obtained from non-inoculated roots and roots synchronized for Glomus intraradices appressorium formation. This approach was conducted in wild-type (J5), mycorrhiza-defective (TRV25, dmi3), and autoregulation-defective (TR122, sunn) M. truncatula genotypes. The groups of proteins that responded to appressorium formation were further compared between wild-type and mutant genotypes; few overlaps and major differences were recorded, demonstrating that mutations in DMI3 and SUNN modified the appressorium-responsive root proteome. Except for a chalcone reductase, none of the differentially displayed proteins that could be identified using matrix-assisted laser desorption ionization time-of-flight mass spectrometry previously was known as appressorium responsive. A DMI3-dependent increased accumulation of signal transduction-related proteins (dehydroascorbate reductase, cyclophilin, and actin depolymerization factor) was found to precede mycorrhiza establishment. Differences in the accumulation of proteins related to plant defense reactions, cytoskeleton rearrangements, and auxin signaling upon symbiont contact were recorded between wild-type and hypermycorrhizal genotypes, pointing to some putative pathways by which SUNN may regulate very early arbuscule formation.
Gene-for-Gene Defense of Wheat Against the Hessian Fly Lacks a Classical Oxidative Burst
Authors: Giovanini Marcelo P. , Puthoff David P. , Nemacheck Jill A. , Mittapalli Omprakash , Saltzmann Kurt D. , Ohm Herbert W. , Shukle Richard H. , Williams Christie E.
Published: May 2007

Genetic similarities between plant interactions with microbial pathogens and wheat interactions with Hessian fly larvae prompted us to investigate defense and counterdefense mechanisms. Plant oxidative burst, a rapid increase in the levels of active oxygen species (AOS) within the initial 24 h of an interaction with pathogens, commonly is associated with defenses that are triggered by gene-for-gene recognition events similar to those involving wheat and Hessian fly larvae. RNAs encoded by Hessian fly superoxide dismutase (SOD) and catalase (CAT) genes, involved in detoxification of AOS, increased in first-instar larvae during both compatible and incompatible interactions. However, mRNA levels of a wheat NADPH oxidase (NOX) gene that generates superoxide (O2¯) did not increase. In addition, inhibiting wheat NOX enzyme with diphenyleneiodonium did not result in increased survival of avirulent larvae. However, nitro blue tetrazolium staining indicated that basal levels of O2¯ are present in both uninfested and infested wheat tissue. mRNA encoded by wheat genes involved in detoxification of the cellular environment, SOD, CAT, and glutathione-S-transferase did not increase in abundance. Histo-chemical staining with 3,3-diaminobenzidine revealed no increases in wheat hydrogen peroxide (H2O2) during infestation that were correlated with the changes in larval SOD and CAT mRNA. However, treatment with 2′,7′-dichlorofluorescin demonstrated the presence of basal levels of H2O2 in the elongation zone of both infested and uninfested plants. The accumulation of a wheat flavanone 3-hydroxylase mRNA did show some parallels with larval gene mRNA profiles. These results suggested that larvae encounter stresses imposed by mechanisms other than an oxidative burst in wheat seedlings.
A Class III Histidine Kinase Acts as a Novel Virulence Factor in Botrytis cinerea
Authors: Viaud Muriel , Fillinger Sabine , Liu Weiwei , Polepalli Jai Santosh , Le Pêcheur Pascal , Kunduru Aditya Reddy , Leroux Pierre , Legendre Laurent
Published: May 2007

Filamentous ascomycetes contain large numbers of histidine kinases (HK) that belong to eleven classes. Members of class III from different species were previously shown to be involved in osmoregulation and resistance to dicarboximide and phenylpyrrole fungicides. We have inactivated the gene encoding the single group III HK, BOS1, in the economically important plant pathogen Botrytis cinerea. BOS1 inactivation had pleiotropic effects on the fungus. Besides the expected osmosensitivity and resistance to fungicides, null mutants presented additional characteristics indicating that BOS1 is necessary for normal macroconidiation and full virulence. On standard culture media, null mutants very rarely formed conidiophores and those few conidiophores failed to produce conidia. This defect could be partially restored with 1 M sorbitol, suggesting that another BOS1-independent signal cascade may be involved in macroconidiation. The mutants were not found to be hypersensitive to various oxidative stresses but were more resistant to menadione. Finally, pathogenicity tests showed that bos1-null mutants were significantly reduced in the ability to infect host plants. Appressorium morphogenesis was not altered; however, in planta growth was severely reduced. To our knowledge, this is the first class III HK characterized as a pathogenicity factor in a plant-pathogenic ascomycete.
WtsE, an AvrE-Family Effector Protein from Pantoea stewartii subsp. stewartii, Causes Disease-Associated Cell Death in Corn and Requires a Chaperone Protein for Stability
Authors: Ham Jong Hyun , Majerczak Doris R. , Arroyo-Rodriguez Angel S. , Mackey David M. , Coplin David L.
Published: May 2007

The pathogenicity of Pantoea stewartii subsp. stewartii to sweet corn and maize requires a Hrp type III secretion system. In this study, we genetically and functionally characterized a disease-specific (Dsp) effector locus, composed of wtsE and wtsF, that is adjacent to the hrp gene cluster. WtsE, a member of the AvrE family of effector proteins, was essential for pathogenesis on corn and was complemented by DspA/E from Erwinia amylovora. An intact C-terminus of WtsE, which contained a putative endoplasmic reticulum membrane retention signal, was important for function of WtsE. Delivery of WtsE into sweet corn leaves by an Escherichia coli strain carrying the hrp cluster of Erwinia chrysanthemi caused water-soaking and necrosis. WtsE-induced cell death was not inhibited by cycloheximide treatment, unlike the hypersensitive response caused by a known Avr protein, AvrRxo1. WtsF, the putative chaperone of WtsE, was not required for secretion of WtsE from P. stewartii, and the virulence of wtsF mutants was reduced only at low inoculum concentrations. However, WtsF was required for full accumulation of WtsE within the bacteria at low temperatures. In contrast, WtsF was needed for efficient delivery of WtsE from E. coli via the Erwinia chrysanthemi Hrp system.
Bioinformatics-Enabled Identification of the HrpL Regulon and Type III Secretion System Effector Proteins of Pseudomonas syringae pv. phaseolicola 1448A
Authors: Vencato Monica , Tian Fang , Alfano James R. , Buell C. Robin , Cartinhour Samuel , DeClerck Genevieve A. , Guttman David S. , Stavrinides John , Joardar Vinita , Lindeberg Magdalen , Bronstein Philip A. , Mansfield John W. , Myers Christopher R. , Collmer Alan , Schneider David J.
Published: May 2007

The ability of Pseudomonas syringae pv. phaseolicola to cause halo blight of bean is dependent on its ability to translocate effector proteins into host cells via the hypersensitive response and pathogenicity (Hrp) type III secretion system (T3SS). To identify genes encoding type III effectors and other potential virulence factors that are regulated by the HrpL alternative sigma factor, we used a hidden Markov model, weight matrix model, and type III targeting-associated patterns to search the genome of P. syringae pv. phaseolicola 1448A, which recently was sequenced to completion. We identified 44 high-probability putative Hrp promoters upstream of genes encoding the core T3SS machinery, 27 candidate effectors and related T3SS substrates, and 10 factors unrelated to the Hrp system. The expression of 13 of these candidate HrpL regulon genes was analyzed by real-time polymerase chain reaction, and all were found to be upregulated by HrpL. Six of the candidate type III effectors were assayed for T3SS-dependent translocation into plant cells using the Bordetella pertussis calmodulin-dependent adenylate cyclase (Cya) translocation reporter, and all were translocated. PSPPH1855 (ApbE-family protein) and PSPPH3759 (alcohol dehydrogenase) have no apparent T3SS-related function; however, they do have homologs in the model strain P. syringae pv. tomato DC3000 (PSPTO2105 and PSPTO0834, respectively) that are similarly upregulated by HrpL. Mutations were constructed in the DC3000 homologs and found to reduce bacterial growth in host Arabidopsis leaves. These results establish the utility of the bioinformatic or candidate gene approach to identifying effectors and other genes relevant to pathogenesis in P. syringae genomes.
The Eight Amino-Acid Differences Within Three Leucine-Rich Repeats Between Pi2 and Piz-t Resistance Proteins Determine the Resistance Specificity to Magnaporthe grisea
Authors: Zhou Bo , Qu Shaohong , Liu Guifu , Dolan Maureen , Sakai Hajime , Lu Guodong , Bellizzi Maria , Wang Guo-Liang
Published: May 2007

The rice blast resistance (R) genes Pi2 and Piz-t confer broad-spectrum resistance against different sets of Magnaporthe grisea isolates. We first identified the Pi2 gene using a map-based cloning strategy. The Pi2 gene is a member of a gene cluster comprising nine gene members (named Nbs1-Pi2 to Nbs9-Pi2) and encodes a protein with a nucleotide-binding site and leucine-rich repeat (LRR) domain. Fine genetic mapping, molecular characterization of the Pi2 susceptible mutants, and complementation tests indicated that Nbs4-Pi2 is the Pi2 gene. The Piz-t gene, a Pi2 allele in the rice cultivar Toride 1, was isolated based on the Pi2 sequence information. Complementation tests confirmed that the family member Nbs4-Piz-t is Piz-t. Sequence comparison revealed that only eight amino-acid changes, which are confined within three consecutive LRR, differentiate Piz-t from Pi2. Of the eight variants, only one locates within the xxLxLxx motif. A reciprocal exchange of the single amino acid between Pi2 and Piz-t did not convert the resistance specificity to each other but, rather, abolished the function of both resistance proteins. These results indicate that the single amino acid in the xxLxLxx motif may be critical for maintaining the recognition surface of Pi2 and Piz-t to their respective avirulence proteins.
The Application of Laser Microdissection to In Planta Gene Expression Profiling of the Maize Anthracnose Stalk Rot Fungus Colletotrichum graminicola
Authors: Tang Weihua , Coughlan Sean , Crane Edmund , Beatty Mary , Duvick Jon
Published: May 2007

Laser microdissection (LM) offers a potential means for deep sampling of a fungal plant-pathogen transcriptome during the infection process using whole-genome DNA microarrays. The use of a fluorescent protein-expressing fungus can greatly facilitate the identification of fungal structures for LM sampling. However, fixation methods that preserve both tissue histology and protein fluorescence, and that also yield RNA of suitable quality for microarray applications, have not been reported. We developed a microwave-accelerated acetone fixation, paraffin-embedding method that fulfills these requirements and used it to prepare mature maize stalk tissues infected with an Anemonia majano cyan fluorescent protein-expressing isolate of the anthracnose stalk rot fungus Colletotrichum graminicola. We successfully used LM to isolate individual maize cells associated with C. graminicola hyphae at an early stage of infection. The LM-derived RNA, after two-round linear amplification, was of sufficient quality and quantity for global expression profiling using a fungal microarray. Comparing replicated LM samples representing an early stage of stalk cell infection with samples from in vitro-germinated conidia, we identified 437 and 370 C. graminicola genes showing significant up- or downregulation, respectively. We confirmed the differential expression of several representative transcripts by quantitative reverse-transcriptase polymerase chain reaction (RT-PCR) and documented extensive overlap of this dataset with a PCR-subtraction library enriched for C. graminicola transcripts in planta. Our results demonstrate that LM is feasible for in planta pathogen expression profiling and can reveal clues about fungal genes involved in pathogenesis. The method in this report may be advantageous for visualizing a variety of cellular features that depend on a high degree of histochemical preservation and RNA integrity prior to LM.
G Protein Signaling Mediates Developmental Processes and Pathogenesis of Alternaria alternata
Authors: Yamagishi Daisuke , Otani Hiroshi , Kodama Motoichiro
Published: May 2007

A G protein α subunit gene (AGA1) has been cloned and characterized from a toxigenic and necrotrophic Alternaria alternata pathogen. Targeted disruption of AGA1 in the apple pathotype of A. alternata gave rise to mutants that differed in colony and conidial morphology as well as sporulation. The conidia of wild type and ΔAGA1 mutants showed equal germination on cellulose membranes. However, wild-type germ tubes formed readily from different points around the conidia, grew randomly, and were often branched, whereas those of the mutants formed only at one or both ends of the conidia and tended to grow in straight paths. Targeted disruption of AGA1 also resulted in reduction of pathogenicity on apple leaves, although the mutant produced host-specific AM-toxin, a fungal secondary metabolite associated with pathogenicity of the pathogen, at levels similar to the wild-type strain. Measurement of the intracellular cAMP levels of the mutant revealed that it was consistently higher than that of the wild type, indicating that AGA1 negatively regulates cAMP levels similar to mammalian Gαi systems. These results indicate that the signal transduction pathway represented by AGA1 appears to be involved in developmental pathways leading to sporulation and pathogenesis of A. alternata.
The Repertoire of Transfer RNA Genes Is Tuned to Codon Usage Bias in the Genomes of Phytophthora sojae and Phytophthora ramorum
Authors: Tripathy Sucheta , Tyler Brett M.
Published: May 2007

In all, 238 and 155 transfer (t)RNA genes were predicted from the genomes of Phytophthora sojae and P. ramorum, respectively. After omitting pseudogenes and undetermined types of tRNA genes, there remained 208 P. sojae tRNA genes and 140 P. ramorum tRNA genes. There were 45 types of tRNA genes, with distinct anticodons, in each species. Fourteen common anticodon types of tRNAs are missing altogether from the genome in the two species; however, these appear to be compensated by wobbling of other tRNA anticodons in a manner which is tied to the codon bias in Phytophthora genes. The most abundant tRNA class was arginine in both P. sojae and P. ramorum. A codon usage table was generated for these two organisms from a total of 9,803,525 codons in P. sojae and 7,496,598 codons in P. ramorum. The most abundant codon type detected from the codon usage tables was GAG (encoding glutamic acid), whereas the most numerous tRNA gene had a me-thionine anticodon (CAT). The correlation between the frequencies of tRNA genes and the codon frequencies in protein-coding genes was very low (0.12 in P. sojae and 0.19 in P. ramorum); however, the correlation between amino acid tRNA gene frequency and the corresponding amino acid codon frequency in P. sojae and P. ramorum was substantially higher (0.53 in P. sojae and 0.77 in P. ramorum). The codon usage frequencies of P. sojae and P. ramorum were very strongly correlated (0.99), as were tRNA gene frequencies (0.77). Approximately 60% of orthologous tRNA gene pairs in P. sojae and P. ramorum are located in regions that have conserved synteny in the two species.
Redox-Active Pyocyanin Secreted by Pseudomonas aeruginosa 7NSK2 Triggers Systemic Resistance to Magnaporthe grisea but Enhances Rhizoctonia solani Susceptibility in Rice
Authors: De Vleesschauwer David , Cornelis Pierre , Höfte Monica
Published: May 2007

Pseudomonas aeruginosa 7NSK2 induces resistance in dicots through a synergistic interaction of the phenazine pyocyanin and the salicylic acid-derivative pyochelin. Root inoculation of the monocot model rice with 7NSK2 partially protected leaves against blast disease (Magnaporthe grisea) but failed to consistently reduce sheath blight (Rhizoctonia solani). Only mutations interfering with pyocyanin production led to a significant decrease in induced systemic resistance (ISR) to M. grisea, and in trans complementation for pyocyanin production restored the ability to elicit ISR. Intriguingly, pyocyanin-deficient mutants, unlike the wild type, triggered ISR against R. solani. Hence, bacterial pyocyanin plays a differential role in 7NSK2-mediated ISR in rice. Application of purified pyocyanin to hydroponically grown rice seedlings increased H2O2 levels locally on the root surface as well as a biphasic H2O2 generation pattern in distal leaves. Co-application of pyocyanin and the antioxidant sodium ascorbate alleviated the opposite effects of pyocyanin on rice blast and sheath blight development, suggesting that the differential effectiveness of pyocyanin with respect to 7NSK2-triggered ISR is mediated by transiently elevated H2O2 levels in planta. The cumulative results suggest that reactive oxygen species act as a double-edged sword in the interaction of rice with the hemibiotroph M. grisea and the necrotroph R. solani.
A Hormone and Proteome Approach to Picturing the Initial Metabolic Events During Plasmodiophora brassicae Infection on Arabidopsis
Authors: Devos Sylvie , Laukens Kris , Deckers Peter , Van Der Straeten Dominique , Beeckman Tom , Inzé Dirk , Van Onckelen Harry , Witters Erwin , Prinsen Els
Published: May 2007

We report on the early response of Arabidopsis thaliana to the obligate biotrophic pathogen Plasmodiophora brassicae at the hormone and proteome level. Using a CYCB1;1::GUS construct, the re-initiation of infection-related cell division is shown from 4 days after inoculation on. Sensitivity to cytokinins and auxins as well as the endogenous hormone levels are evaluated. Both an enhanced cytokinin gene response and an accumulation of isopentenyl adenine and adenosine precede this re-initiation of cell division, whereas an enhanced auxin gene response is observed from 6 days after inoculation on. The alh1 mutant, impaired in the cross talk between ethylene and auxins, is resistant to P. brassicae. A differential protein analysis of infected versus noninfected roots and hypocotyls was performed using two-dimensional gel electrophoresis and quantitative image analysis, coupled to matrix-assisted laser desorption ionization time of flight-time of flight mass spectrometry-based protein identification. Of the visualized proteins, 12% show altered abundance compared with the nonin-fected plants, including proteins involved in metabolism, cell defense, cell differentiation, and detoxification. Combining the hormone and proteome data, we postulate that, at the very first stages of Plasmodiophora infection, plasmodial-produced cytokinins trigger a local re-initiation of cell division in the root cortex. Consequently, a de novo meristematic area is established that acts as a sink for host-derived indole-3-acetic acid, carbohydrates, nitrogen, and energy to maintain the pathogen and to trigger gall development.
The Pathogen-Host Interactions Database (PHI-base) Provides Insights into Generic and Novel Themes of Pathogenicity
Authors: Baldwin Thomas K. , Winnenburg Rainer , Urban Martin , Rawlings Chris , Koehler Jacob , Hammond-Kosack Kim E.
Published: May 2007

Fungal and oomycete pathogens of plants and animals are a major global problem. In the last 15 years, many genes required for pathogenesis have been determined for over 50 different species. Other studies have characterized effector genes (previously termed avirulence genes) required to activate host responses. By studying these types of pathogen genes, novel targets for control can be revealed. In this report, we describe the Pathogen-Host Interactions database (PHI-base), which systematically compiles such pathogenicity genes involved in pathogen-host interactions. Here, we focus on the biology that underlies this computational resource: the nature of pathogen-host interactions, the experimental methods that exist for the characterization of such pathogen-host interactions as well as the available computational resources. Based on the data, we review and analyze the specific functions of pathogenicity genes, the host-specific nature of pathogenicity and virulence genes, and the generic mechanisms of effectors that trigger plant responses. We further discuss the utilization of PHI-base for the computational identification of pathogenicity genes through comparative genomics. In this context, the importance of standardizing pathogenicity assays as well as integrating databases to aid comparative genomics is discussed.
Requirement of a mip-Like Gene for Virulence in the Phytopathogenic Bacterium Xanthomonas campestris pv. campestris
Authors: Zang Ning , Tang Dong-Jie , Wei Mei-Liang , He Yong-Qiang , Chen Baoshan , Feng Jia-Xun , Xu Jing , Gan Yong-Qi , Jiang Bo-Le , Tang Ji-Liang
Published: May 2007

Macrophage infectivity potentiators (Mips) are FKBP domain-containing proteins reported as virulence factors in several human pathogens, such as members of genera Legionella, Salmonella and Chlamydia. The putative peptidyl-prolyl cis-trans isomerase (PPIase) encoded by XC2699 of the plant bacterial pathogen Xanthomonas campestris pv. campestris 8004 exhibits a 49% similarity at the aminoacid level to the Mip protein of Legionella pneumophila. This mip-like gene, XC2699, was overexpressed in Es-cherichia coli and the purified (His)6-tagged Mip-like protein encoded by XC2699 exhibited a PPIase activity specifically inhibited by FK-506. A mutation in the mip-like gene XC2699 led to significant reductions in virulence and replication capacity in the host plant Chinese radish (Raphanus sativus L. var. radiculus Pers.). Furthermore, the production of exopolysaccharide and the activity of extracellular proteases, virulence factors X. campestris pv. campestris, were significantly decreased in the mip-like mutant. These results reveal that the mip-like gene is involved in the pathogenesis of X. campestris pv. campestris through an effect on the production of these virulence factors.
Functional Interplay Between Two Xanthomonas oryzae pv. oryzae Secretion Systems in Modulating Virulence on Rice
Authors: Jha Gopaljee , Rajeshwari Ramanan , Sonti Ramesh V.
Published: May 2007

The type II (T2S) and type III (T3S) secretion systems are important for virulence of Xanthomonas oryzae pv. oryzae, causal agent of bacterial leaf blight of rice. The T3S of gram-negative bacterial plant pathogens has been shown to suppress host defense responses, including programmed cell death reactions, whereas the T2S is involved in secreting cell-wall-degrading enzymes. Here, we show that a T3S-deficient (T3S¯) mutant of X. oryzae pv. oryzae can induce a basal plant defense response seen as callose deposition, immunize rice against subsequent X. oryzae pv. oryzae infection, and cause cell-death-associated nuclear fragmentation. A T2S¯ T3S¯ double mutant exhibited a substantial reduction in the ability to evoke these responses. We purified two major effectors of the X. oryzae pv. oryzae T2S and characterized them to be a cellulase (ClsA) and a putative cellobiosidase (CbsA). The purified ClsA, CbsA, and lipase/esterase (LipA; a previously identified T2S effector) proteins induced rice defense responses that were suppressible by X. oryzae pv. oryzae in a T3S-dependent manner. These defense responses also were inducible by the products of the action of these purified proteins on rice cell walls. We further show that a CbsA¯ mutant or a ClsA¯ LipA¯ double mutant are severely virulence deficient. These results indicate that the X. oryzae pv. oryzae T2S secretes important virulence factors, which induce innate rice defense responses that are suppressed by T3S effectors to enable successful infection.
Nod Factor Perception During Infection Thread Growth Fine-Tunes Nodulation
Authors: Den Herder Jeroen , Vanhee Celine , De Rycke Riet , Corich Viviana , Holsters Marcelle , Goormachtig Sofie
Published: May 2007

Bacterial nodulation factors (NFs) are essential signaling molecules for the initiation of a nitrogen-fixing symbiosis in legumes. NFs are perceived by the plant and trigger both local and distant responses, such as curling of root hairs and cortical cell divisions. In addition to their requirement at the start, NFs are produced by bacteria that reside within infection threads. To analyze the role of NFs at later infection stages, several phases of nodulation were studied by detailed light and electron microscopy after coinoculation of adventitious root primordia of Sesbania rostrata with a mixture of Azorhizobium caulinodans mutants ORS571-V44 and ORS571-X15. These mutants are deficient in NF production or surface polysaccharide synthesis, respectively, but they can complement each other, resulting in functional nodules occupied by ORS571-V44. The lack of NFs within the infection threads was confirmed by the absence of expression of an early NF-induced marker, leghemoglobin 6 of S. rostrata. NF production within the infection threads is shown to be necessary for proper infection thread growth and for synchronization of nodule formation with bacterial invasion. However, local production of NFs by bacteria that are taken up by the plant cells at the stage of bacteroid formation is not required for correct symbiosome development.
Transcriptional Adaptation of Mycosphaerella graminicola to Programmed Cell Death (PCD) of Its Susceptible Wheat Host
Authors: Keon John , Antoniw John , Carzaniga Raffaella , Deller Siân , Ward Jane L. , Baker John M. , Beale Michael H. , Hammond-Kosack Kim , Rudd Jason J.
Published: May 2007

Many important fungal pathogens of plants spend long periods (days to weeks) of their infection cycle in symptomless association with living host tissue, followed by a sudden transition to necrotrophic feeding as host tissue death occurs. Little is known about either the host responses associated with this sudden transition or the specific adaptations made by the pathogen to invoke or tolerate it. We are studying a major host-specific fungal pathogen of cultivated wheat, Septoria tritici (teleomorph Mycosphaerella graminicola). Here, we describe the host responses of wheat leaves infected with M. graminicola during the development of disease symptoms and use microarray transcription profiling to identify adaptive responses of the fungus to its changing environment. We show that symptom development on a susceptible host genotype has features reminiscent of the hypersensitive response, a rapid and strictly localized form of host programmed cell death (PCD) more commonly associated with disease-resistance mechanisms. The initiation and advancement of this host response is associated with a loss of cell-membrane integrity and dramatic increases in apoplastic metabolites and the rate of fungal growth. Micro-array analysis of the fungal genes differentially expressed before and after the onset of host PCD supports a transition to more rapid growth. Specific physiological adaptation of the fungus is also revealed with respect to membrane transport, chemical and oxidative stress mechanisms, and metabolism. Our data support the hypothesis that host plant PCD plays an important role in susceptibility towards fungal pathogens with necrotrophic lifestyles.
Two-Component Sensor RhpS Promotes Induction of Pseudomonas syringae Type III Secretion System by Repressing Negative Regulator RhpR
Authors: Xiao Yanmei , Lan Lefu , Yin Chuntao , Deng Xin , Baker Douglas , Zhou Jian-Min , Tang Xiaoyan
Published: May 2007

The Pseudomonas syringae type III secretion system (T3SS) is induced during interaction with the plant or culture in minimal medium (MM). How the bacterium senses these environments to activate the T3SS is poorly understood. Here, we report the identification of a novel two-component system (TCS), RhpRS, that regulates the induction of P. syringae T3SS genes. The rhpR and rhpS genes are organized in an operon with rhpR encoding a putative TCS response regulator and rhpS encoding a putative biphasic sensor kinase. Transposon insertion in rhpS severely reduced the induction of P. syringae T3SS genes in the plant as well as in MM and significantly compromised the pathogenicity on host plants and hypersensitive response-inducing activity on nonhost plants. However, deletion of the rhpRS locus allowed the induction of T3SS genes to the same level as in the wild-type strain and the recovery of pathogenicity upon infiltration into plants. Overexpression of RhpR in the ΔrhpRS deletion strain abolished the induction of T3SS genes. However, overexpression of RhpR in the wild-type strain or overexpression of RhpR(D70A), a mutant of the predicted phosphorylation site of RhpR, in the ΔrhpRS deletion strain only slightly reduced the induction of T3SS genes. Based on these results, we propose that the phosphorylated RhpR represses the induction of T3SS genes and that RhpS reverses phosphorylation of RhpR under the T3SS-inducing conditions. Epistasis analysis indicated that rhpS and rhpR act upstream of hrpR to regulate T3SS genes.
Single-Cell Transcript Profiling of Barley Attacked by the Powdery Mildew Fungus
Authors: Gjetting Torben , Hagedorn Peter H. , Schweizer Patrick , Thordal-Christensen Hans , Carver Timothy L. W. , Lyngkjær Michael F.
Published: May 2007

In many plant-pathogen interactions, there are several possible outcomes for simultaneous attacks on the same leaf. For instance, an attack by the powdery mildew fungus on one barley leaf epidermal cell may succeed in infection and formation of a functional haustorium, whereas a neighboring cell attacked at the same time may resist fungal penetration. To date, the mixed cellular responses seen even in susceptible host leaves have made it difficult to relate induced changes in gene expression to resistance or susceptibility in bulk leaf samples. By microextraction of cell-specific mRNA and subsequent cDNA array analysis, we have successfully obtained separate gene expression profiles for specific mildew-resistant and -infected barley cells. Thus, for the first time, it is possible to identify genes that are specifically regulated in infected cells and, presumably, involved in fungal establishment. Further, although much is understood about the genetic basis of effective papilla resistance associated with mutant mlo barley, we provide here the first evidence for gene regulation associated with effective papilla-based nonspecific resistance expressed in nominally “susceptible” wild-type barley.
Characterization of Genomic Clones and Expression Analysis of the Three Types of Superoxide Dismutases During Nodule Development in Lotus japonicus
Authors: Rubio Maria C. , Becana Manuel , Sato Shusei , James Euan K. , Tabata Satoshi , Spaink Herman P.
Published: May 2007

Superoxide dismutases (SODs) are metalloenzymes that play a primary role in the protection against oxidative stress in plants and other organisms. We have characterized four SOD genes in Lotus japonicus and have analyzed their expression in roots and four developmental stages of nodules. The expression of cytosolic CuZnSOD, at the mRNA, protein, and enzyme activity levels, decreases with nodule age, and the protein is localized in the dividing cells and infection threads of emergent nodules and in the infected cells of young nodules. The mitochondrial MnSOD was downregulated, whereas the bacteroidal MnSOD displayed maximal protein and enzyme activity levels in older nodules. Two additional genes, encoding plastidic (FeSOD1) and cytosolic (FeSOD2) FeSOD isoforms, were identified and mapped. The genes are located in different chromosomes and show differential expression. The FeSOD1 mRNA level did not change during nodule development, whereas FeSOD2 was upregulated. The distinct expression patterns of the SOD genes may reflect different regulatory mechanisms of the enzyme activities during nodule ontogeny. In particular, at the mRNA and activity levels, the virtual loss of cytosolic CuZnSOD in mature and old nodules, concomitant with the induction of FeSOD2, suggests that the two enzymes may functionally compensate each other in the cytosol at the late stages of nodule development.
Coi1-Dependent Signaling Pathway Is Not Required for Mi-1—Mediated Potato Aphid Resistance
Authors: Bhattarai Kishor K. , Xie Qi-Guang , Pourshalimi Daniel , Younglove Ted , Kaloshian Isgouhi
Published: May 2007

Tomato (Solanum lycopersicum) has a unique resistance gene, Mi-1, that confers resistance to animals from distinct taxa, nematodes, and piercing and sucking insects. Mi-1 encodes a protein with a nucleotide-binding site and leucine-rich repeat motifs. Early in the potato aphid (Macrosiphum euphorbiae)—tomato interactions, aphid feeding induces the expression of the jasmonic acid (JA)-regulated proteinase inhibitor genes, Pin1 and Pin2. The jai1-1 (jasmonic acid insensitive 1) tomato mutant, which is impaired in JA perception, was used to gain additional insight into the JA signaling pathway and its role in the Mi-1—mediated aphid resistance. The jai1-1 mutant has a deletion in the Coi1 gene that encodes a putative F-box protein. In this study, aphid colonization, survival, and fecundity were compared on wild-type tomato and jai1-1 mutant. In choice assays, the jai1-1 mutant showed higher colonization by potato aphids compared with wild-type tomato. In contrast, no-choice assays showed no difference in potato aphid survival or fecundity between jai1-1 and the wild-type parent. Plants homozygous for Mi-1 and for the jai1 mutation were not compromised in resistance to potato aphids, using either choice or no-choice assays. In addition, the accumulation of JA-regulated Pin1 transcripts after aphid feeding was Coi1 dependent. Taken together, these data indicate that, although potato aphids activate Coi1-dependent defense response in tomato, this response is not required for Mi-1—mediated resistance to aphids.
Parallel Genome-Wide Expression Profiling of Host and Pathogen During Soybean Cyst Nematode Infection of Soybean
Authors: Ithal Nagabhushana , Recknor Justin , Nettleton Dan , Hearne Leonard , Maier Tom , Baum Thomas J. , Mitchum Melissa G.
Published: May 2007

Global analysis of gene expression changes in soybean (Glycine max) and Heterodera glycines (soybean cyst nematode [SCN]) during the course of infection in a compatible interaction was performed using the Affymetrix GeneChip soybean genome array. Among 35,611 soybean transcripts monitored, we identified 429 genes that showed statistically significant differential expression between uninfected and nematode-infected root tissues. These included genes encoding enzymes involved in primary metabolism; biosynthesis of phenolic compounds, lignin, and flavonoids; genes related to stress and defense responses; cell wall modification; cellular signaling; and transcriptional regulation. Among 7,431 SCN transcripts monitored, 1,850 genes showed statistically significant differential expression across different stages of nematode parasitism and development. Differentially expressed SCN genes were grouped into nine different clusters based on their expression profiles during parasitism of soybean roots. The patterns of gene expression we observed in SCN suggest coordinated regulation of genes involved in parasitism. Quantitative real-time reverse-transcription polymerase chain reaction confirmed the results of our microarray analysis. The simultaneous genome-wide analysis of gene expression changes in the host and pathogen during a compatible interaction provides new insights into soybean responses to nematode infection and the first profile of transcript abundance changes occurring in the nematode as it infects and establishes a permanent feeding site within a host plant root.
The Role of a P1-Type ATPase from Pseudomonas fluorescens SBW25 in Copper Homeostasis and Plant Colonization
Authors: Zhang Xue-Xian , Rainey Paul B.
Published: May 2007

The genome of the plant-colonizing bacterium Pseudomonas fluorescens SBW25 possesses a putative copper-transporting P1-type ATPase (CueA) that is induced on the plant surfaces. Using a chromosomally-integrated cueA-'lacZ fusion, we show that transcription of cueA can be induced (in vitro) by ions of copper, silver, gold, and mercury. To investigate the biological significance of cueA, a nonpolar cueA deletion mutant (SBW25ΔcueA) was constructed. This mutant strain displayed a twofold reduction in its tolerance to copper compared with the wild-type strain; however, no change was observed in the sensitivity of the mutant strain to silver, gold, or mercury ions. To obtain insight into the ecological significance of cueA, the competitive ability of SBW25ΔcueA was determined relative to wild-type SBW25 in three environments (none contained added copper): minimal M9 medium, the root of sugar beet (Beta vulgaris), and the root of pea (Pisum sativum). Results showed that the fitness of SBW25ΔcueA was not different from the wild type in laboratory medium but was compromised in the two plant environments. Taken together, these data demonstrate a functional role for CueA in copper homeostasis and reveal an ecologically significant contribution to bacterial fitness in the plant rhizosphere. They also suggest that copper ions accumulate on plant surfaces.
Role of the K-Antigen Subgroup of Capsular Polysaccharides in the Early Recognition Process Between Rhizobium meliloti and Alfalfa Leaves
Authors: Becquart-de Kozak Isabelle , Reuhs Bradley L. , Buffard Dominique , Breda Colette , Kim John S. , Esnault Robert , Kondorosi Adam
Published: May 2007

We used a model system to investigate the induction of plant genes by bacterial surface compounds. We have infiltrated alfalfa leaves with wild-type Rhizobium meliloti strain Rm41 and mutant derivatives, which are deficient in the production of exopolysaccharides (EPS), capsular K-polysaccharides (KPS or K-Antigens), or both. We have shown that wild-type R. meliloti was able to induce transcript accumulation of genes encoding enzymes of the iso-flavonoid biosynthetic pathway: chalcone synthase (CHS), chalcone reductase (CHR), and isoflavone reductase (IFR). Kinetics of these transcript accumulations were biphasic, occurring 0.75 to 1.5 h and 6 to 30 h after treatment. The exoB derivative of Rm41 (strain AK631), which is unable to produce either EPS I or EPS II, was still able to induce very rapid (45-min posttreatment) transcript accumulation of CHS and CHR genes. These results suggested that EPS were not involved in this induction. We then tested three R. meliloti fix-23 mutants (strains PP699, PP711, and PP671), which were deficient in KPS production as shown by PAGE and immunoblot analyses. In contrast to the results obtained with the mutant strains, infiltration of alfalfa leaves with purified KPS, isolated from strain AK631, led to a very rapid induction of the CHS and CHR genes. Therefore, we hypothesize that KPS may play a role in the early recognition of rhizobia by the leaf cells.
Classical and Molecular Genetics of the Model Legume Lotus japonicus
Authors: Jiang Qunyi , Gresshoff Peter M.
Published: May 2007

The model legume Lotus japonicus was demonstrated to be amenable to classical and molecular genetic analysis, providing the basis for the genetic dissection of the plant processes underlying nodulation and nitrogen fixation. We have developed an efficient method for the sexual hybridization of L. japonicus and obtained F1 progeny derived from a cross of L. japonicus B-129-S9 Gifu × B-581 Funakura. Over half of the cross-pollinations resulted in fertile hybrid seed, which were confirmed morphologically and by single arbitrary primer DNA amplification polymorphisms using the DAF technique. Molecular and morphological markers segregated in true Mendelian fashion in a F2 population of 100 plants. Several DAF loci were linked using the MAPMAKER software to create the first molecular linkage groups of this model legume. The mapping population was advanced to generate a set of immortal recombinant inbred lines (F6; RILs), useful for sharing plant material fixed genetically at most genomic regions. Morphological loci for waved stem shape (Ssh), dark leaf color (Lco), and short flowering period (Fpe) were inherited as single dominant Mendelian loci. DAF markers were dominant and were detected between Gifu and Funakura at about one per primer, suggesting that the parents are closely related. One polymorphism (270G generated by single octomer primer 8.6m) was linked to a morphological locus controlling leaf coloration. The results demonstrate that (i) Lotus japonicus is amenable to diploid genetic analysis, (ii) morphological and molecular markers segregate in true diploid fashion, (iii) molecular polymorphisms can be obtained at a reasonable frequency between the related Gifu and Funakura lines, and iv) the possibility exists for map-based cloning, marker assisted selection and mapping of symbiotic mutations through a genetic and molecular map.