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Pierre Rouge - One of the best experts on this subject based on the ideXlab platform.
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a molecular basis for the endo β1 3 glucanase activity of the thaumatin like proteins from edible fruits
Biochimie, 2003Co-Authors: Laurence Menubouaouiche, Christelle Vriet, Willy J Peumans, Annick Barre, Els J M Van Damme, Pierre RougeAbstract:Abstract Fruit-specific thaumatin-like proteins were isolated from cherry, apple and banana, and their enzymatic and antifungal activities compared. Both the apple and cherry possess a moderate endo-β1,3-glucanase activity but are devoid of antifugal activity. In contrast, the banana thaumatin-like protein inhibits the in vitro hyphal growth of Verticillium Albo-Atrum but is virtually devoid of endo-β1,3-glucanase activity. Both structural and molecular modeling studies showed that all three thaumatin-like proteins possess an extended electronegatively charged cleft at their surface, which is believed to be a prerequisite for endo-β1,3-glucanase activity. Docking experiments showed that the positioning of linear (1,3)-β-D-glucans in the cleft of the apple and cherry proteins allows an interaction with the glutamic acid residues that are responsible for the hydrolytic cleavage of the glucan. Due to a different positioning in the cleft of the banana thaumatin-like protein, the linear β-glucans cannot properly interact with the catalytic glutamic acid residues and as a result the protein possesses no enzymatic activity. The possible function of the fruit-specific thaumatin-like proteins is discussed in view of the observed biological activities and structural features.
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a molecular basis for the endo β1 3 glucanase activity of the thaumatin like proteins from edible fruits
Biochimie, 2003Co-Authors: Laurence Menubouaouiche, Els J M Van Damme, Christelle Vriet, Willy J Peumans, Annick Barre, Pierre RougeAbstract:Fruit-specific thaumatin-like proteins were isolated from cherry, apple and banana, and their enzymatic and antifungal activities compared. Both the apple and cherry possess a moderate endo-beta 1,3-glucanase activity but are devoid of antifugal activity. In contrast, the banana thaumatin-like protein inhibits the in vitro hyphal growth of Verticillium Albo-Atrum but is virtually devoid of endo-beta 1,3-glucanase activity. Both structural and molecular modeling studies showed that all three thaumatin-like proteins possess an extended electronegatively charged cleft at their surface, which is believed to be a prerequisite for endo-beta 1,3-glucanase activity. Docking experiments showed that the positioning of linear (1,3)-beta-D-glucans in the cleft of the apple and cherry proteins allows an interaction with the glutamic acid residues that are responsible for the hydrolytic cleavage of the glucan. Due to a different positioning in the cleft of the banana thaumatin-like protein, the linear beta-glucans cannot properly interact with the catalytic glutamic acid residues and as a result the protein possesses no enzymatic activity. The possible function of the fruit-specific thaumatin-like proteins is discussed in view of the observed biological activities and structural features.
Branka Javornik - One of the best experts on this subject based on the ideXlab platform.
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Evaluation of reference genes for RT-qPCR expression studies in hop (Humulus lupulus L.) during infection with vascular pathogen Verticillium Albo-Atrum
2016Co-Authors: Sara Cregeen, Branka JavornikAbstract:Hop plant (Humulus lupulus L.), cultivated primarily for its use in the brewing industry, is faced with a variety of diseases, including severe vascular diseases, such as Verticillium wilt, against which no effective protection is available. The understanding of disease resistance with tools such as differentially expressed gene studies is an important objective of plant defense mechanisms. In this study, we evaluated twenty-three reference genes for RT-qPCR expression studies on hop under biotic stress conditions. The candidate genes were validated on susceptible and resistant hop cultivars sampled at three different time points after infection with Verticillium Albo-Atrum. The stability of expression and the number of genes required for accurate normalization were assessed by three different Excel-based approaches (geNorm v.3.5 software, NormFinder, and RefFinder). High consistency was found among them, identifying the same six best reference genes (YLS8
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the secretome of vascular wilt pathogen Verticillium albo atrum in simulated xylem fluid
Proteomics, 2015Co-Authors: Stanislav Mandelc, Branka JavornikAbstract:Verticillium Albo-Atrum is a vascular wilt pathogen capable of infecting many important dicotyledonous plant species. Fungal isolates from hop differ in aggressiveness, causing either mild or lethal symptoms in infected plants. As in other plant pathogenic fungi, extracellular proteins, such as cell wall-degrading enzymes and effectors, are thought to be crucial in the pathogenesis process. In this study, mild and lethal isolates from three countries were grown in simulated xylem medium and secretome analysis by 2D-DIGE showed low qualitative and high quantitative variability among the isolates. Functional classification of 194 identified proteins representing 100 unique protein accessions revealed an arsenal of cell wall-degrading enzymes and potential effectors. The set of proteins that were more abundant in at least two lethal isolates included enzymes acetylcholinesterases, lipases, polygalacturonases, pectate lyase, rhamnogalacturonan acetylesterases, acetylxylan esterase, endoglucanase, xylanases, mannosidases, and a protein similar to alginate lyase and also potential effectors necrosis- and ethylene-inducing protein, small basic 14 kDa hypothetical protein and 79 kDa hypothetical proteins. Other proteins associated with virulence showed different expression profiles between mild and lethal isolates. The results suggest that the increased virulence of lethal isolates has little background shared by all three lethal isolates and that upregulation of isolate specific sets of proteins may be most important.
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ORIGINAL PAPER Different Gene Expressions of Resistant and Susceptible Hop Cultivars in Response to Infection with a Highly Aggressive Strain
2014Co-Authors: Sara Cregeen, Sebastjan Radišek, Natasa Stajner, Stanislav M, Boris Turk, Jernej Jakse, Branka JavornikAbstract:# The Author(s) 2014. This article is published with open access at Springerlink.com Abstract Verticillium wilt has become a serious threat to hop production in Europe due to outbreaks of lethal wilt caused by a highly virulent strain of Verticillium Albo-Atrum. In order to enhance our understanding of resistance mechanisms, the fungal colonization patterns and interactions of resistant and susceptible hop cultivars infected with V. Albo-Atrum were analysed in time course experiments. Quantification of fungal DNA showed marked differences in spatial and temporal fungal colonization patterns in the two cultivars. Two differ-ential display methods obtained 217 transcripts with altered expression, of which 84 showed similarity to plant proteins and 8 to fungal proteins. Gene ontology categorised them into cellular and metabolic processes, response to stimuli, biolog-ical regulation, biogenesis and localization. The expression patterns of 17 transcripts with possible implication in plant immunity were examined by real-time PCR (RT-qPCR). Our results showed strong expression of genes encoding pathogenesis-related (PR) proteins in susceptible plants and strong upregulation of genes implicated in ubiquitination and vesicle trafficking in the incompatible interaction and their downregulation in susceptible plants, suggesting the involve-ment of these processes in the hop resistance reaction. In the resistant cultivar, the RT-qPCR expression patterns of most genes showed their peak at 20 dpi and declined towards 30 dpi, comparable to the gene expression pattern of in planta detected fungal protein and coinciding with the highest fungal biomass in plants at 15 dpi. These expression patterns suggest that the defence response in the resistant cultivar is strong enough at 20 dpi to restrict further fungus colonization
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evaluation of reference genes for rt qpcr expression studies in hop humulus lupulus l during infection with vascular pathogen Verticillium albo atrum
PLOS ONE, 2013Co-Authors: Natasa Stajner, Sara Cregeen, Branka JavornikAbstract:Hop plant (Humulus lupulus L.), cultivated primarily for its use in the brewing industry, is faced with a variety of diseases, including severe vascular diseases, such as Verticillium wilt, against which no effective protection is available. The understanding of disease resistance with tools such as differentially expressed gene studies is an important objective of plant defense mechanisms. In this study, we evaluated twenty-three reference genes for RT-qPCR expression studies on hop under biotic stress conditions. The candidate genes were validated on susceptible and resistant hop cultivars sampled at three different time points after infection with Verticillium Albo-Atrum. The stability of expression and the number of genes required for accurate normalization were assessed by three different Excel-based approaches (geNorm v.3.5 software, NormFinder, and RefFinder). High consistency was found among them, identifying the same six best reference genes (YLS8, DRH1, TIP41, CAC, POAC and SAND) and five least stably expressed genes (CYCL, UBQ11, POACT, GAPDH and NADH). The candidate genes in different experimental subsets/conditions resulted in different rankings. A combination of the two best reference genes, YLS8 and DRH1, was used for normalization of RT-qPCR data of the gene of interest (PR-1) implicated in biotic stress of hop. We outlined the differences between normalized and non-normalized values and the importance of RT-qPCR data normalization. The high correlation obtained among data standardized with different sets of reference genes confirms the suitability of the reference genes selected for normalization. Lower correlations between normalized and non-normalized data may reflect different quantity and/or quality of RNA samples used in RT-qPCR analyses.
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Average expression stability values (M) calculated by geNorm.
2013Co-Authors: Natasa Stajner, Sara Cregeen, Branka JavornikAbstract:The genes are ranked according to their expression stability; the least stable genes are on the left, and the two most stable genes are on the right. The samples are divided into sub-groups: (A) susceptible plants of cv. Celeia; (B) resistant plants of cv. Wye Target; (C) plants infected with Verticillium Albo-Atrum; (D) control (non-infected) plants; (E) plants evaluated 10 days post inoculation with fungi; (F) plants evaluated 20 days post inoculation with fungi (G) plants evaluated 30 days post inoculation with fungi (H) all plant samples.
Laurence Menubouaouiche - One of the best experts on this subject based on the ideXlab platform.
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a molecular basis for the endo β1 3 glucanase activity of the thaumatin like proteins from edible fruits
Biochimie, 2003Co-Authors: Laurence Menubouaouiche, Christelle Vriet, Willy J Peumans, Annick Barre, Els J M Van Damme, Pierre RougeAbstract:Abstract Fruit-specific thaumatin-like proteins were isolated from cherry, apple and banana, and their enzymatic and antifungal activities compared. Both the apple and cherry possess a moderate endo-β1,3-glucanase activity but are devoid of antifugal activity. In contrast, the banana thaumatin-like protein inhibits the in vitro hyphal growth of Verticillium Albo-Atrum but is virtually devoid of endo-β1,3-glucanase activity. Both structural and molecular modeling studies showed that all three thaumatin-like proteins possess an extended electronegatively charged cleft at their surface, which is believed to be a prerequisite for endo-β1,3-glucanase activity. Docking experiments showed that the positioning of linear (1,3)-β-D-glucans in the cleft of the apple and cherry proteins allows an interaction with the glutamic acid residues that are responsible for the hydrolytic cleavage of the glucan. Due to a different positioning in the cleft of the banana thaumatin-like protein, the linear β-glucans cannot properly interact with the catalytic glutamic acid residues and as a result the protein possesses no enzymatic activity. The possible function of the fruit-specific thaumatin-like proteins is discussed in view of the observed biological activities and structural features.
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a molecular basis for the endo β1 3 glucanase activity of the thaumatin like proteins from edible fruits
Biochimie, 2003Co-Authors: Laurence Menubouaouiche, Els J M Van Damme, Christelle Vriet, Willy J Peumans, Annick Barre, Pierre RougeAbstract:Fruit-specific thaumatin-like proteins were isolated from cherry, apple and banana, and their enzymatic and antifungal activities compared. Both the apple and cherry possess a moderate endo-beta 1,3-glucanase activity but are devoid of antifugal activity. In contrast, the banana thaumatin-like protein inhibits the in vitro hyphal growth of Verticillium Albo-Atrum but is virtually devoid of endo-beta 1,3-glucanase activity. Both structural and molecular modeling studies showed that all three thaumatin-like proteins possess an extended electronegatively charged cleft at their surface, which is believed to be a prerequisite for endo-beta 1,3-glucanase activity. Docking experiments showed that the positioning of linear (1,3)-beta-D-glucans in the cleft of the apple and cherry proteins allows an interaction with the glutamic acid residues that are responsible for the hydrolytic cleavage of the glucan. Due to a different positioning in the cleft of the banana thaumatin-like protein, the linear beta-glucans cannot properly interact with the catalytic glutamic acid residues and as a result the protein possesses no enzymatic activity. The possible function of the fruit-specific thaumatin-like proteins is discussed in view of the observed biological activities and structural features.
Bart P. H. J. Thomma - One of the best experts on this subject based on the ideXlab platform.
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Verticillium alfalfae and V . dahliae , Agents of Verticillium Wilt Diseases
Genomics of Plant-Associated Fungi and Oomycetes: Dicot Pathogens, 2014Co-Authors: Patrik Inderbitzin, Bart P. H. J. Thomma, S. J. Klosterman, Krishna V SubbaraoAbstract:Verticillium dahliae and V. alfalfae (formerly Verticillium Albo-Atrum) are two important agricultural pathogens that affect many crops around the world and cause a distinct type of vascular wilt, which are known as Verticillium wilts. Several V. alfalfae and V. dahliae genomes have been sequenced, and are among the smaller genomes from filamentous ascomycetes. The number of predicted protein-encoding genes is similar to the saprobe Neurospora crassa. Perhaps reflective of their particular hemibiotrophic life styles, some gene families are expanded in the V. alfalfae and V. dahliae genomes. These include the gene families encoding glycoside hydrolases GH88, necrosis and ethylene-inducing-like proteins (NLPs), LysM effectors, proteins with chitin-recognition motifs, and cutinases. But the number of predicted secreted proteins was less than half that of the related Colletotrichum species, the agents of anthracnose diseases. V. dahliae strains generally contain lineage-specific regions (LS regions), which may play an important role in virulence and pathogenicity. Examples for horizontal transfer into Verticillium ancestors include the virulence factor Ave1, a glucan glucosyltransferase, and potentially some of the retrotransposons. The V. alfalfae and V. dahliae genomes have already had significant impacts on various aspects of basic and applied Verticillium research.
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interfamily transfer of tomato ve1 mediates Verticillium resistance in arabidopsis
Plant Physiology, 2011Co-Authors: Emilie F Fradin, Ahmed Abdelhaliem, Laura Masini, G C M Van Den Berg, M H A J Joosten, Bart P. H. J. ThommaAbstract:Vascular wilts caused by soil-borne fungal species of the Verticillium genus are devastating plant diseases. The most common species, Verticillium dahliae and Verticillium Albo-Atrum, have broad host ranges and are notoriously difficult to control. Therefore, genetic resistance is the preferred method for disease control. Only from tomato (Solanum lycopersicum) has a Verticillium resistance locus been cloned, comprising the Ve1 gene that encodes a receptor-like protein-type cell surface receptor. Due to lack of a suitable model for receptor-like protein (RLP)-mediated resistance signaling in Arabidopsis (Arabidopsis thaliana), so far relatively little is known about RLP signaling in pathogen resistance. Here, we show that Ve1 remains fully functional after interfamily transfer to Arabidopsis and that Ve1-transgenic Arabidopsis is resistant to race 1 but not to race 2 strains of V. dahliae and V. Albo-Atrum, nor to the Brassicaceae-specific pathogen Verticillium longisporum. Furthermore, we show that signaling components utilized by Ve1 in Arabidopsis to establish Verticillium resistance overlap with those required in tomato and include SERK3/BAK1, EDS1, and NDR1, which strongly suggests that critical components for resistance signaling are conserved. We subsequently investigated the requirement of SERK family members for Ve1 resistance in Arabidopsis, revealing that SERK1 is required in addition to SERK3/BAK1. Using virus-induced gene silencing, the requirement of SERK1 for Ve1-mediated resistance was confirmed in tomato. Moreover, we show the requirement of SERK1 for resistance against the foliar fungal pathogen Cladosporium fulvum mediated by the RLP Cf-4. Our results demonstrate that Arabidopsis can be used as model to unravel the genetics of Ve1-mediated resistance.
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design and development of a dna array for rapid detection and identification of multiple tomato vascular wilt pathogens
Fems Microbiology Letters, 2003Co-Authors: Bart Lievens, M H J Brouwer, A C R C Vanachter, Andre C Levesque, Bruno P A Cammue, Bart P. H. J. ThommaAbstract:Fusarium wilt, caused by Fusarium oxysporum f. sp. lycopersici, and Verticillium wilt, caused by either Verticillium Albo-Atrum or Verticillium dahliae, are devastating diseases of tomato (Lycopersicon esculentum) found worldwide. Monitoring is the cornerstone of integrated pest management of any disease. The lack of rapid, accurate, and reliable means by which plant pathogens can be detected and identified is one of the main limitations in integrated disease management. In this paper, we describe the development of a molecular detection system, based on DNA array technology, for rapid and efficient detection of these vascular wilt pathogens. We show the utility of this array for the sensitive detection of these pathogens from complex substrates like soil, plant tissues and irrigation water, and samples that are collected by tomato growers in their greenhouses.
Annick Barre - One of the best experts on this subject based on the ideXlab platform.
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a molecular basis for the endo β1 3 glucanase activity of the thaumatin like proteins from edible fruits
Biochimie, 2003Co-Authors: Laurence Menubouaouiche, Christelle Vriet, Willy J Peumans, Annick Barre, Els J M Van Damme, Pierre RougeAbstract:Abstract Fruit-specific thaumatin-like proteins were isolated from cherry, apple and banana, and their enzymatic and antifungal activities compared. Both the apple and cherry possess a moderate endo-β1,3-glucanase activity but are devoid of antifugal activity. In contrast, the banana thaumatin-like protein inhibits the in vitro hyphal growth of Verticillium Albo-Atrum but is virtually devoid of endo-β1,3-glucanase activity. Both structural and molecular modeling studies showed that all three thaumatin-like proteins possess an extended electronegatively charged cleft at their surface, which is believed to be a prerequisite for endo-β1,3-glucanase activity. Docking experiments showed that the positioning of linear (1,3)-β-D-glucans in the cleft of the apple and cherry proteins allows an interaction with the glutamic acid residues that are responsible for the hydrolytic cleavage of the glucan. Due to a different positioning in the cleft of the banana thaumatin-like protein, the linear β-glucans cannot properly interact with the catalytic glutamic acid residues and as a result the protein possesses no enzymatic activity. The possible function of the fruit-specific thaumatin-like proteins is discussed in view of the observed biological activities and structural features.
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a molecular basis for the endo β1 3 glucanase activity of the thaumatin like proteins from edible fruits
Biochimie, 2003Co-Authors: Laurence Menubouaouiche, Els J M Van Damme, Christelle Vriet, Willy J Peumans, Annick Barre, Pierre RougeAbstract:Fruit-specific thaumatin-like proteins were isolated from cherry, apple and banana, and their enzymatic and antifungal activities compared. Both the apple and cherry possess a moderate endo-beta 1,3-glucanase activity but are devoid of antifugal activity. In contrast, the banana thaumatin-like protein inhibits the in vitro hyphal growth of Verticillium Albo-Atrum but is virtually devoid of endo-beta 1,3-glucanase activity. Both structural and molecular modeling studies showed that all three thaumatin-like proteins possess an extended electronegatively charged cleft at their surface, which is believed to be a prerequisite for endo-beta 1,3-glucanase activity. Docking experiments showed that the positioning of linear (1,3)-beta-D-glucans in the cleft of the apple and cherry proteins allows an interaction with the glutamic acid residues that are responsible for the hydrolytic cleavage of the glucan. Due to a different positioning in the cleft of the banana thaumatin-like protein, the linear beta-glucans cannot properly interact with the catalytic glutamic acid residues and as a result the protein possesses no enzymatic activity. The possible function of the fruit-specific thaumatin-like proteins is discussed in view of the observed biological activities and structural features.
