Academic literature on the topic 'Hypersensitive response (HR)'

Strains of Clavibacter michiganensis subsp. sepedonicus, causal agent of bacterial ring rot of potato, showed marked differences in virulence on host plants. When infiltrated into tobacco leaves, virulent strains caused a rapid localized necrotic response (within 24 to 48 h) characteristic of the hypersensitive response (HR), whereas nonpathogenic strains did not. Concentrated cell-free culture supernatants (CCS) from virulent strains caused a necrotic reaction on tobacco, whereas CCS from nonpathogenic strains did not. The necrosis-inducing activity was heat stable and protease sensitive. Inhibitors of eukaryotic metabolism suppressed the necrotic reaction of tobacco to CCS. No necrotic response was observed when host plants were infiltrated with either cells or CCS from virulent strains. HR-inducing protein(s) from a virulent strain separated from the majority of other proteins on DEAE cellulose at 250 to 300 mM NaCl. Ammonium sulfate-precipitated proteins from a virulent strain produced a necrotic reaction at a total protein concentration of 18 μg/ml, whereas those from a nonpathogenic strain did not, even at a concentration of 180 μg/ml. We conclude that virulent strains of C. michiganensis subsp. sepedonicus elicit a typical HR in tobacco and secrete proteinaceous elicitor(s) of the nonhost HR.

The plant hypersensitive response (HR) to avirulent bacterial pathogens results from programmed cell death of plant cells in the infected region. Ion leakage and changes in signaling components associated with HR progression were measured. These studies compared Arabidopsis mutants affecting feedback loops with wild-type plants, with timepoints taken hourly. In response to Pseudomonas syringae pv. tomato DC3000·avrB, npr1-2 mutant plants showed increased ion leakage relative to wild-type plants. Hydrogen peroxide accumulation was similar to that in wild type, but salicylic acid accumulation was reduced at some timepoints. With DC3000·avrRpt2, similar trends were seen. In response to DC3000·avrB, ndr1-1 mutant plants showed more ion leakage than wild-type or npr1-2 plants. Hydrogen peroxide accumulation was delayed by approximately 1 h and reached half the level seen with wild-type plants. Salicylic acid accumulation was similar to npr1-2 mutant plants. With DC3000·avrRpt2, ndr1-1 mutant plants showed no ion leakage, no hydrogen peroxide accumulation, and minimal salicylic acid accumulation. Results with a ndr1-1 and npr1-2 double mutant were similar to ndr1-1. A model consistent with these data is presented, in which one positive and two negative regulatory circuits control HR progression. Understanding this circuitry will facilitate HR manipulation for enhanced disease resistance.

Tobacco infected with Pseudomonas syringae pv. phaseolicola undergoes a hypersensitive response (HR). Jasmonic acid (JA) accumulated within the developing lesion 3 to 9 h after infection and this accumulation preceded protein loss, cell death, and malondialdehyde accumulation. Accumulating JA consisted largely of the (—)-JA stereoisomer and was essentially restricted to the HR lesion.

The Pca crown rust resistance cluster in the diploid Avena genus confers gene-for-gene specificity to numerous isolates of Puccinia coronata f. sp. avenae. Recombination breakpoint analysis indicates that specificities conferred by the Pca cluster are controlled by at least five distinct genes, designated Pc81, Pc82, Pc83, Pc84, and Pc85. Avena plants with the appropriate genotype frequently respond to P. coronata by undergoing hypersensitive cell death at the sites of fungal infection. Autofluorescence of host cells in response to P. coronata occurs in plants that develop visible necrotic lesions but not in plants that lack this phenotype. Two newly described, non-Pc loci were shown to control hypersensitive cell death. Rds (resistance-dependent suppressor of cell death) suppresses the hypersensitive response (HR), but not the resistance, mediated by the Pc82 resistance gene. In contrast, Rih (resistance-independent hypersensitive cell death) confers HR in both resistant and susceptible plants. Linkage analysis indicates that Rds is unlinked to the Pca cluster, whereas Rih is tightly linked to it. These results indicate that multiple synchronous pathways affect the development of hypersensitive cell death and that HR is not essential for resistance to crown rust. Further characterization of these genes will clarify the relationship between plant disease resistance and localized hypersensitive cell death.

In tobacco plants reacting hypersensitively to pathogen infection, localized acquired resistance (LAR) is induced in a sharp zone surrounding hypersensitive response (HR) lesions. Using a fungal glycoprotein inducing HR and LAR when infiltrated at 50 nM into tobacco leaves, we have shown previously that a plant signal(s) is released by HR cells and diffuses to induce LAR. Here we address two questions: does LAR occur when HR is not induced, and is salicylic acid the (or one of the) mobile LAR signal? We found that application to tobacco leaves of 0.25 nM glycoprotein triggered defense responses without HR and without an H2O2 burst. The analyzed responses include changes in expression of O-methyltransferase (OMT), 3-hydroxy-3-methylglutarylCoA reductase, pathogenesis-related (PR) proteins, and changes in levels of the signal salicylic acid. No defense responses and no increased resistance to tobacco mosaic virus infection were found beyond the elicitor-infiltrated tissue, providing strong evidence that there is no LAR without HR. Treatments of NahG tobacco leaves with 50 nM elicitor induced the HR and, in the sharp zone surrounding the HR lesion, a strong activation of OMT and of basic PR proteins, but not of acidic PR-1 proteins. This indicates that a signal different from salicylic acid is diffusing.

Solanum melongena (eggplant) exhibits a hypersensitive response (HR) when infected with tobacco mosaic tobamovirus (TMV). In contrast, a TMV mutant unable to express coat protein (CP) did not elicit the HR, while a potexvirus vector engineered to express TMV CP did elicit the eggplant HR. The CPs of U2 and odontoglossum ringspot tobamoviruses also elicited the HR. However, the HR was not elicited by the CP of cucumber green mottle mosaic tobamovirus. Taken together, these findings demonstrate that specific tobamovirus CPs function as elicitors of the eggplant HR.

One of the most studied defense reactions of plants against microbial pathogens is the hypersensitive response (HR). The HR is a complex multicellular process that involves programmed cell death at the site of infection. A standard method to quantify plant defense and the HR is to measure the release of cellular electrolytes into water after infiltration with pathogenic bacteria. In this type of experiment, the bacteria are typically delivered into the plant tissue through syringe infiltration. Here we report the development of a vacuum infiltration protocol that allows multiple plant lines to be infiltrated simultaneously and assayed for defense responses. Vacuum infiltration did not induce more wounding response in Arabidopsis leaf tissue than syringe inoculation, whereas throughput and reproducibility were improved. The method was used to study HR-induced electrolyte loss after treatment with the bacteriumPseudomonas syringaepv.tomatoDC3000 harboring the effector AvrRpm1, AvrRpt2 or AvrRps4. Specifically, the influence of bacterial titer on AvrRpm1-induced HR was investigated. Not only the amplitude, but also the timing of the maximum rate of the HR reaction was found to be dose-dependent. Finally, using vacuum infiltration, we were able quantify induction of phospholipase D activity after AvrRpm1 recognition in leaves labeled with33PO4.

Rsv1, a single dominant gene in soybean PI 96983, confers resistance to most strains of Soybean mosaic virus (SMV), including strain G2. The phenotypic response includes the lack of symptoms and virus recovery from mechanically inoculated leaves. To study the resistance mechanism, SMV-N (an isolate of strain G2) was introduced into PI 96983 by grafting. Hypersensitive response (HR)-like lesions occurred on the stems, petioles, and leaf veins, and virus was recovered from these lesions. The response demonstrated the cytological and histological characteristics of HR as well as elevated transcription of a soybean salicylic acid-inducible, pathogenesis-related (PR-1) protein gene. Mechanical inoculation of PI 96983 primary leaves with a high level of SMV-N virions caused no symptoms or up regulation of the PR-1 protein gene transcript. Furthermore, inoculation with infectious viral RNA did not alter the resistance phenotype. The data suggest that interaction of SMV-N with Rsv1 has the potential to induce an HR-like defense reaction. Rsv1-mediated resistance in the inoculated leaf, however, is HR-independent and operates after virion disassembly.

The plant hypersensitive response (HR) is tightly associated with gene-for-gene resistance and has been proposed to function in containing pathogens at the invasion site. This tight association has made it difficult to unequivocally evaluate the importance of HR for plant disease resistance. Here, hopPsyA from Pseudomonas syringae pv. syringae 61 is identified as a new avirulence gene for Arabidopsis that triggers resistance in the absence of macroscopic HR. Resistance to P. syringae pv. tomato DC3000 expressing hopPsyA was EDS1-dependent and NDR1-independent. Intriguingly, several Arabidopsis accessions were resistant to DC3000(hopPsyA) in the absence of HR. This is comparable to the Arabidopsis response to avrRps4, but it is shown that hopPsyA does not signal through RPS4. In a cross between two hopPsyA-resistant accessions that differ in their HR response, the HR segregated as a recessive phenotype regulated by a single locus. This locus, HED1 (HR regulator in EDS1 pathway), is proposed to encode a protein whose activity can cause suppression of the EDS1-dependent HR signaling pathway. HED1-regulated symptomless gene-for-gene resistance responses may explain some cases of Arabidopsis resistance to bacteria that are classified as nonhost resistance.

The objective of this study was to transform sweet pepper ferredoxin-like protein (PFLP) gene, which has antimicrobial properties, to tobacco and investigate the disease resistance abilities of transgenic tobacco. This protein interacts with another protein, harpin that is produced by the bacteria which is invading the plant tissues, and stimulates hypersensitivity response in plants, thus the spreading of disease is limited. Gene transfer was achieved to tobacco by Agrobacterium- mediated method and with indirect organogenesis<br>the explants were grown on selective media and then transferred to jars and pots respectively. Molecular and genetic analyses such as PCR, RT-PCR, Sequence Analysis and Northern Blot, were performed with plants which their seeds survived and grew on selective medium and also gave positive reactions for GUS histochemical assay. Finally, with putative transgenic plants, some hypersensitive response assays were carried out with Pseudomonas syringae and it was observed that the recovered plants showed hypersensitive response (HR) in the preliminary tests. These results indicated that putative transgenic tobacco plants which carry pflp transgene, can be used in disease resistance studies.

Diversiteten bland gener som ger resistens (R) mot infektionssjukdomar är mycket stor i växtriket, särskilt i den effektorutlösta klassen av försvaret. Denna artikel beskriver interaktioner mellan växters effektorutlösta fösvar och patogeners effektorproteiner och undersöker evolutionära och genetiska mekanismer för uppkomsten av ny resistens. R-genprodukter har en typisk domänstruktur och interagerar både direkt och indirekt med patogeners effektorproteiner.Vid kontakt med ett effektorprotein utlöses en försvarsrespons som kan förhindra fortsatt patogen tillväxt. Flera genetiska mekanismer verkar samtidigt på R-gener, något som resulterar i en hög hastighet för uppkomst av nya R-varianter. Den intima samevolution som existerar hos många patogen-växtsystem formar det evolutionära utvecklingsmönstret av R-gener. Diversifierande positiv selektion via den biologiska kapprustningsmodellen och bevarande negativ frekvensberoende selektion är båda viktiga samevolutionära processer. Denna artikel indikerar att växters immunförsvar är anpassningsbart och robust, men belyser också den kunskapsbrist som råder inom resistensforskningen.<br>The diversity among genes conferring resistance (R) against infectious diseases is very high in the plant kingdom, especially in the effector triggered class of defense. This article describes the interactions between the plant effector triggered immunity and pathogen effectors and examines the evolutionary and genetic mechanisms for the emergence of new resistance. R gene products have a typical domain structure and interacts both directly and indirectly with pathogen effectors. Upon contact with an effector a defense response is triggered that may prevent the further pathogen growth. Multiple genetic mechanisms act simultaneously on the R-genes, which results in a rapid diversification of novel R variants. The intimate co-evolution of many existing plant-pathogen systems form the evolutionary pattern of R genes. Diversifying positive selection via the biological arms race model and conservative negative frequency-dependent selection are both important coevolutionary processes. This article indicates that the plant immune system is adaptive and robust, but also highlights the lack of knowledge in plant resistance research.

L'ossido nitrico (NO) è una molecola gassosa radicale e una molecola di segnalazione chiave coinvolto nella mediazione di varie condizioni di sviluppo e di risposta allo stress biotico e abiotico nelle piante. Nell'interazione pianta-patogeno, NO presenta un ruolo cruciale nella difesa contro i microbi biotrofici, lavorando in collaborazione con ROS per innescare la morte cellulare nella zona infetta; un meccanismo noto come risposta ipersensibile (HR). Tuttavia, i meccanismi molecolari di come NO coordina questo processo è ancora sconosciuta. Per identificare i geni coinvolti nella segnalazione NO durante il processo di morte cellulare ipersensibile, abbiamo effettuato uno screening genetico diretto, utilizzando un sistema di doppia selezione per identificare piante mutanti di Arabidopsis compromessi nella signaling di NO durante la morte ipersensibile. Per lo screening primario, una piattaforma di fumigazione di NO è stato sviluppato e in seguito è stato identificato condizioni per innescare la morte cellulare in piante wild-type di Arabidopsis indotta dal trattamento col NO. Fumigando in totale 25.600 M2 piante provenienti dal etil metano sulfonato (EMS) e della popolazione mutante con neutroni (FN), abbiamo identificato 19 linee mutanti compromessi nella morte cellulare indotta dal NO. Il secondo screening consisteva nell'esaminare la riduzione della HR indotta dal patogeno in questi mutanti pre-selezionati. Analizzando 13 linee mutanti, resistenti al’NO, per una diminuzione della morte cellulare innescata dell'infezione con patogeni non virulenti, abbiamo identificato 7 linee mutanti che sono stati alterati in questo processo. Le caratterizzazione dei processi riguardo alla HR è stata eseguita in tre di questi mutanti selezionati. Analizzando il loro sistema antiossidante e la presenza di normale produzione di NO e ROS durante la risposta ipersensibile in queste mutanti è stato possibile allocare la loro mutazione in posizione diversa nella segnalazione di NO durante la risposta di resistenza delle piante. I nostri risultati hanno dimostrato che i tre mutanti mostravano alterazioni nel sistema antiossidante, ciò influenzato negativamente la produzioni di ROS in uno degli mutanti. Inoltre, la sotto regolazione dell'attività della catalase ha contribuito per il loro superiore livello di ROS. A quanto riguarda la produzione di NO, questo è stato ridotto invece in tutte le tre linee ma, solo una linea ha presentato una completamente compromessa produzione di NO. È interessante notare che, in tutti i tre mutanti è stato disturbato la modulazione dell'attività GSNOR durante l’HR. Concludendo, i nostri dati indicano che non solo il livello di produzione di entrambe NO e ROS, ma anche i meccanismi per la loro rimozioni sono stati determinanti per avere il normale sviluppo della morte ipersensibile; e quindi, la strategia di selezione progettato potrebbe anche selezionare mutanti deteriorati in questi processi. In questo modo, questi risultati rinforzano l'importanza di avere biodisponibilità equilibrata tra H2O2 e NO nell'esecuzione della morte cellulare programmata nella risposta di resistenza delle piante al patogeno. La caratterizzazione della produzione di NO e di ROS in altri mutanti identificati permetterà di selezionare mutanti più interessanti da sequenziare. La preparazione della popolazione di mappatura utilizzando piante mutanti M3 originali ad incroci (OC) si ha rivelato di essere negativo per il nostro studio. La presenza di genotipi polimorfici nella popolazione ibrido F2 OC ha prodotto una segregazione genetica distorta, precludendo per determinare in modo inequivocabile il gene mutato. Pertanto, strategia supplementare per l'identificazione della mutazione causale, come l'uso di mutanti F2 backcrossed per la creazione di una popolazione di mappatura, dovrebbe essere considerata una volta che saranno prescelti candidati mutanti migliori.<br>Nitric oxide (NO) is a gaseous free radical and a key signaling molecule involved in the mediation of various developmental and stress-related conditions in plants. In plant-pathogen interaction, NO plays a crucial role in defense against biotrophic microbes, working together with ROS to trigger cell death in the infected area; a mechanism known as hypersensitive response (HR). However, the molecular mechanisms of how NO coordinates this process is still unknown. To identify candidate genes involved in NO signaling during the HR-cell death process, we carried out a forward genetic screen, using a double-selection system to identify Arabidopsis mutant plants compromised in NO-mediated HR-cell death. For the primary screening, an NO fumigation platform was developed and conditions inducing uniform cell death on wild-type Arabidopsis plants identified. Fumigating in total 25,600 M2 ethyl methane sulfonate (EMS) and fast neutron (FN) mutant plants, we identified 19 mutant lines as non-responsive for NO-induced cell death. The second screening step consisted then, in examining the reduction in pathogen induced HR-cell death in these pre-selected mutants. Evaluation of 13 NO resistant mutant lines for decreased cell death upon avirulent pathogen infection, we finally identified 7 mutant lines that were impaired in this process. An extensive characterization of HR-cell death process was performed in three of these selected mutants. Analyzing the antioxidant system and the presence of normal NO and ROS bursts in these lines, was possible to allocate their mutation in different position in NO signaling in plant resistance. Our results demonstrated that all three mutants showed alteration in the antioxidant system, what affected negatively the ROS burst in one of them. The down-regulation of catalase activity contributed for their higher endogenous level of ROS. Moreover, the NO burst was reduced instead in all three lines but, only one presented a fully compromised NO burst. Interestingly, in all three mutants the modulation of GSNOR activity during HR was disturbed. Concluding, our data indicated that not only the rate of NO and ROS production but also the mechanisms for their turnover were determinant for having normal development of HR-cell death; and therefore, the designed selection strategy also could select mutants impaired in these processes. Thus, these findings reinforce the importance of having balanced bioavailability between H2O2 and NO in the execution of cell death program in plant resistance response. Characterization of NO and ROS in other identified mutants will allow to select most interesting mutants to be sequenced. Mapping population obtained from original M3 mutant plants for outcrossing (OC) revealed to be not the best material for our study. The presence of polymorphic genotypes in the hybrid F2 OC population yielded a distorted genetic segregation and could preclude to determine unequivocally, the mutated gene. Therefore additional strategy for identification of the causal mutation, like the use of F2 backcrossed mutants for creating a mapping population, should be considered once best candidates will be finally selected.

The gram-negative bacterium Pseudomonas syringae pv. actinidiae (Psa) is the causal agent of the bacterial canker of kiwifruits leading to important economic losses in New Zealand, Italy, Korea and Japan the main producers worldwide. To identify virulence targets and develop new effective targeted control strategies, molecular mechanisms involved in host plant recognition by Psa and related infection processes was currently studied. To this aim, the Psa CRAFRU 8.43 strain was transformed with constructs carrying the green fluorescent protein (GFP)-encoding reporter gene under the control of the promoter of several Psa genes, selected for their putative role in pathogenicity based on literature. Our results showed that hrpA1 gene promoter, involved in the early steps of bacterial infection, is induced in a minimal medium (mimicking apoplast conditions), with an earlier and at higher levels in presence of Actinidia deliciosa leaf extract, indicating a role downstream of host recognition by bacteria. The characterization of hrpA1-inducing kiwi extract showed that such signal(s) are kiwi-specific and smaller than 10KDa in size. Moreover, to elucidate the signalling pathway(s) involved in host-mediated hrpA1 induction, a chemical library was screened to identify molecules able to block such activation. Nineteen candidate molecules were obtained, displaying different inhibition levels. According to the role of HrpA1 in plant hypersensitive response (HR) induction, HR in model plants wasquantifiedto obtain a visible phenotype correlated with hrpA1 promoter induction and thus demonstrate the inhibitory effect of selected molecule, dicoumarol, in a plant system. Moreover, the molecules responsible for Psa quorum sensing activation are still unknown, since Psa possess three LuxR-solos proteins (PsaR1, R2, R3) lacking LuxI enzymes responsible for AHL synthesis. Thus, to identify putative PsaR1 ligands, its recombinant autoinducer-binding domain was produced in E. coli and chemical libraries were screened using a high throughput fluorescence-based thermal shift assay. Four molecules inducing a significant thermal shift were identified as putative PsaR1 ligands; moreover, the presence of the same or other putative PsaR1 ligands in kiwi plants was confirmed by a similar thermal shift effect in presence of kiwi plant extracts. Quercetin and luteolin, PsaR1 putative ligands, tested on Psa virulence traits (motility and biofilm) showed an increase of swarming motility and reduction biofilm formation.

Infrared thermography is used to reveal the establishment of Erwinia amylovora harpin-induced hypersensitive response (HR) in Nicotiana sylvestris leaves. We observed a decrease in temperature (1-2°C) in the harpin infiltrated zone, correlated with an increase in stomatal opening, strongly suggesting that the temperature decrease is due to higher transpiration rate. IRT experiments were conducted in a laboratory environment and could be widely applied for genotype screening and monitoring drug effects.

Plant-microbe interactions involve a large number of global regulatory systems, which are essential for plants to protect themselves against pathogen attack. An ethylene-inducing xylanase (EIX) of Trichoderma viride is a potent elicitor of plant defense responses, like hypersensitive response (HR), in specific cultivars of tobacco (Nicotiana tabacum) and tomato (Lycopersicon esculentum). The central goal of this proposal was to investigate the molecular mechanisms that allow plants to specifically activate defense responses after EIX treatment. We proposed to identify cellular signaling components involved in the induction of HR by the EIX elicitor. The molecular genetic analysis of the signal transduction pathway that modulates hypersensitive responses is an important step in understanding the induction of plant defense responses. The genes that mediate LeEIX2-EIX dependent activation of resistance mechanisms remain to be identified. We used two approaches to identify the cellular signaling components that induce HR mediated by the EIX elicitor. In the first approach, we performed a yeast two-hybrid screening using LeEix2 as bait to identify plant proteins that interact with it. In the second approach, we used virus-induced gene silencing (VIGS) for a high-throughput screen to identify genes that are required for the induction of LeEIX2-EIX mediated HR. VIGS will also be used for functional characterization of genes that will be identified during the yeast two-hybrid screen. This investigation will shed light on cellular processes and signaling components involved in induction of general plant defense against pathogens and will provide the basis for future biotechnological approaches to improve plant resistance to pathogens. Several genes were indentified by the two approaches. We used the VIGS and yeast two hybrid approaches to confirm that activity of the genes initially identified by different procedure. Two genes inhibit the induction of HR by the fungal elicitor in the different systems; Tobacco-Harpin binding protein 1 and cyclopropyl isomerase.