Virtual Issue Articles: Host surface properties affect prepenetration processes in the barley powdery mildew fungus Vanessa Zabka et al
Plant signalling components EDS1 and SGT1 enhance disease caused by the necrotrophic pathogen Botrytis cinerea Mohamed El Oirdi and Kamal Bouarab
Evolutionary history of the ancient cutinase family in five filamentous Ascomycetes reveals differential gene duplications and losses and in Magnaporthe grisea shows evidence of suband neo-functionalization Pari Skamnioti, Rebecca F. Furlong and Sarah J. Gurr
Tracks for traffic: mirotubules in the plant pathogen Ustilago maydis Gero Steinberg
Effectors of biotrophic fungi and oomycetes: pathogenicity factors and triggers of host resistance Peter N. Dodds et al Magnaporthe grisea avirulence gene ACE1 belongs to an infection-specific gene cluster involved in secondary metabolism Jérôme Collemare et al Early and specific gene expression triggered by rice resistance gene Pi33 in response to infection by ACE1 avirulent blast fungus E. Vergne, E. Ballini et al The Lmgpi15 gene, encoding a component of the glycosylphosphatidylinositol anchor biosynthesis pathway, is required for morphogenesis and pathogenicity in Leptosphaeria maculans Estelle Remy et al Fusarium graminearum gene deletion mutants map1 and tri5 reveal similarities and differences in the pathogenicity requirements to cause disease on Arabidopsis and wheat floral tissue Alayne Cuzick, Martin Urban and Kim Hammond-Kosack Fusarium graminearum exploits ethylene signalling to colonize dicotyledonous and monocotyledonous plants X. Chen, A. Steed, S. Travella, B. Keller and P. Nicholson
Microbiological control of soil-borne phytopathogenic fungi with special emphasis on wilt-inducing Fusarium oxysporum Claude Alabouvette, Chantal Olivain, Quirico Migheli and Christian Steinberg The use of FLP-mediated recombination for the functional analysis of an effector gene family in the biotrophic smut fungus Ustilago maydis Yuliya Khrunyk et al Dynamic carbon transfer during pathogenesis of sunflower by the necrotrophic fungus Botrytis cinerea: from plant hexoses to mannitol Thierry Dulermo et al Expression profiling and functional analysis of Populus WRKY23 reveals a regulatory role in defense Valérie Levée et al Natural genetic resources of Arabidopsis thaliana reveal a high prevalence and unexpected phenotypic plasticity of RPW8mediated powdery mildew resistance Katharina Göllner, Patrick Schweizer, Yuling Bai and Ralph Panstruga RPW8 and resistance to powdery mildew pathogens in natural populations of Arabidopsis lyrata T. H. Jorgensen and B. C. Emerson
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Virtual Issue Articles: Activation tagging of ADR2 conveys a spreading lesion phenotype and resistance to biotrophic pathogens Mourad A. M. Aboul-Soud et al
Phytotoxic Nep1-like proteins from the necrotrophic fungus Botrytis cinerea associate with membranes and the nucleus of plant cells Alexander Schouten, Peter Van Baarlen and Jan A. L. Van Kan
Specificity and levels of nonhost resistance to nonadapted Blumeria graminis forms in barley Reza Aghnoum and Rients E. Niks Role of hydrogen peroxide during the interaction between the hemibiotrophic fungal pathogen Septoria tritici and wheat Nandini P. Shetty et al
Cover image from: Aghnoum and Niks (2009). v.185, pp. 275-284.
De novo biosynthesis of defense root exudates in response to Fusarium attack in barley Arnaud Lanoue et al Action and reaction of host and pathogen during Fusarium head blight disease Stephanie Walter, Paul Nicholson and Fiona M. Doohan Arabidopsis thaliana plant defensin AtPDF1.1 is involved in the plant response to biotic stress Barbara M. A. De Coninck et al Tête à tête inside a plant cell: establishing compatibility between plants and biotrophic fungi and oomycetes Richard J. O’Connell and Ralph Panstruga Frozen in time: a new method using cryo-scanning electron microscopy to visualize root–fungal interactions Steve Refshauge, Michelle Watt, Margaret E. McCully and Cheng X. Huang Multivesicular compartments proliferate in susceptible and resistant MLA12-barley leaves in response to infection by the biotrophic powdery mildew fungus Qianli An et al
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Introduction Plants are continuously threatened by a plethora of biotic stresses that include encounters with viruses, bacteria, fungi, nematodes and herbivorous animals. Amongst these threats, diseases caused by fungal pathogens have a particular impact, accounting for major damage and yield losses in agriculture. Basic research on such plant diseases is thus pivotal to advance our understanding of molecular mechanisms underlying fungal pathogenesis and plant defence. In the long term, these activities may lead to rational strategies for durable disease control that will result in a reduction in fungicide usage. New Phytologist has a long-standing tradition in the publication of original research articles and reviews that address molecular, physiological, evolutionary and environmental aspects of symbiotic and pathogenic plant–fungus interactions. In this Virtual Special Issue we focus on ‘Pathogenic plant–fungus interactions’ and have compiled a selection of papers recently published in New Phytologist that include research on the topics of molecular mechanisms of fungal pathogenesis, plant defence signalling and response, and analyses of the plant–fungus interface.
Host surface properties affect prepenetration processes in the barley powdery mildew fungus Vanessa Zabka, Michaela Stangl, Gerhard Bringmann, Gerd Vogg, Markus Riederer and Ulrich Hildebrandt
Summary Author for correspondence: Ulrich Hildebrandt Tel: +49 (0)931 888 6206 Fax: +49 (0)931 888 6235 Email: ulrich.hildebrandt@botanik. uni-wuerzburg.de
KEYWORDS: Blumeria graminis; barley (Hordeum vulgare); hexacosanal; hexacosanol; hydrophobicity; mildew; wax
The initial contact between Blumeria graminis f.sp. hordei and its host barley (Hordeum vulgare) takes place on epicuticular waxes at the surfaces of aerial plant organs. Here, the extent to which chemical composition, crystal structure and hydrophobicity of cuticular waxes affect fungal prepenetration processes was explored.
The leaf surface properties of barley eceriferum (cer) wax mutants were characterized in detail. Barley leaves and artificial surfaces were used to investigate the early events of fungal infection.
Even after epicuticular waxes had been stripped away, cer mutant leaf surfaces did not affect fungal prepenetration properties. Removal of total leaf cuticular waxes, however, resulted in a 20% reduction in conidial germination and differentiation. Two major components of barley leaf wax, hexacosanol and hexacosanal, differed considerably in their ability to effectively trigger conidial differentiation on glass surfaces. While hexacosanol, attaining a maximum hydrophobicity with contact angles of no more than 80, proved to be noninductive, hexacosanal significantly stimulated differentiation in c. 50% of B. graminis conidia, but only at contact angles > 80.
These results, together with an observed inductive effect of highly hydrophobic, wax-free artificial surfaces, provide new insights into the interplay of physical and chemical surface cues involved in triggering prepenetration processes in B. graminis.
Evolutionary history of the ancient cutinase family in five filamentous Ascomycetes reveals differential gene duplications and losses and in Magnaporthe grisea shows evidence of sub- and neo-functionalization Pari Skamnioti, Rebecca F. Furlong and Sarah J. Gurr
Author for correspondence: Pari Skamnioti and Sarah J. Gurr Tel: +44 1865 275116 Fax: +44 1865 275074 Emails: pari.skamnioti@plants.ox.ac.uk (PS) sarah.gurr@plants.ox.ac.uk (SJG)
Summary
The cuticle is the first barrier for fungi that parasitize plants systematically or opportunistically. Here, the evolutionary history is reported of the multimembered cutinase families of the plant pathogenic Ascomycetes Magnaporthe grisea, Fusarium graminearum and Botrytis cinerea and the saprotrophic Ascomycetes Aspergillus nidulans and Neurospora crassa.
New Phytologist (2008) 180: 711–721 doi: 10.1111/j.1469-8137.2008.02598.x
Molecular taxonomy of all fungal cutinases demonstrates a clear division into two ancient subfamilies. No evidence was found for lateral gene transfer from prokaryotes. The cutinases in the five Ascomycetes show significant copy number variation, they form six clades and their extreme sequence diversity is highlighted by the lack of consensus intron. The average ratio of gene duplication to loss is 2 : 3, with the exception of M. grisea and N. crassa, which exhibit extreme family expansion and contraction, respectively.
Detailed transcript profiling in vivo, categorizes the M. grisea cutinases into four regulatory patterns. Symmetric or asymmetric expression profiles of phylogenetically related cutinase genes suggest subfunctionalization and neofunctionalization, respectively.
The cutinase family-size per fungal species is discussed in relation to genome characteristics and lifestyle. The ancestry of the cutinase gene family, together with the expression divergence of its individual members provides a first insight into the drivers for niche differentiation in fungi.
KEYWORDS: Ascomycetes, duplication, evolution, expression analysis, gene family, Magnaporthe, neofunctionalization, subfunctionalization.
Effectors of biotrophic fungi and oomycetes: pathogenicity factors and triggers of host resistance Peter N. Dodds, Maryam Rafiqi, Pamela H. P. Gan, Adrienne R. Hardham, David A. Jones and Jeffrey G. Ellis
Summary Author for correspondence: Peter N. Dodds Tel: +61 2 6246 5039 Email: peter.dodds@csiro.au
New Phytologist (2009) 183: 993–1000 doi: 10.1111/j.1469-8137.2009.02922.x
KEYWORDS: avirulence, biotroph, effector proteins, haustoria, oomycete, resistance, rust.
Many biotrophic fungal and oomycete pathogens share a common infection process involving the formation of haustoria, which penetrate host cell walls and form a close association with plant membranes. Recent studies have identified a class of pathogenicity effector proteins from these pathogens that is transferred into host cells from haustoria during infection. This insight stemmed from the identification of avirulence (Avr) proteins from these pathogens that are recognized by intracellular host resistance (R) proteins. Oomycete effectors contain a conserved translocation motif that directs their uptake into host cells independently of the pathogen, and is shared with the human malaria pathogen. Genome sequence information indicates that oomycetes may express several hundred such host-translocated effectors. Elucidating the transport mechanism of fungal and oomycete effectors and their roles in disease offers new opportunities to understand how these pathogens are able to manipulate host cells to establish a parasitic relationship and to develop new disease-control measures.
Magnaporthe grisea avirulence gene ACE1 belongs to an infectionspecific gene cluster involved in secondary metabolism Jérôme Collemare, Mikaël Pianfetti, Anne-Elodie Houlle, Damien Morin, Laurent Camborde, MarieJosèphe Gagey, Crystel Barbisan, Isabelle Fudal, Marc-Henri Lebrun and Heidi U. Böhnert
Summary Author for correspondence: Marc-Henri Lebrun Tel: +33 4 72 85 24 81 Fax: +33 4 72 85 22 97 Email:
The avirulence gene ACE1 from the rice blast fungus Magnaporthe grisea encodes a polyketide synthase (PKS) fused to a nonribosomal peptide synthetase (NRPS) probably involved in the biosynthesis of a secondary metabolite recognized by Pi33 resistant rice (Oryza sativa) cultivars.
marc-henri.lebrun@bayercropscience.com
New Phytologist (2008) 179: 196–208 doi: 10.1111/j.1469-8137.2008.02459.x
KEYWORDS: appressorium, expression pattern, PKS-NRPS, regulatory network, rice blast fungus (Magnaporthe grisea).
Analysis of the M. grisea genome revealed that ACE1 is located in a cluster of 15 genes, of which 14 are potentially involved in secondary metabolism as they encode enzymes such as a second PKS-NRPS (SYN2), two enoyl reductases (RAP1 and RAP2) and a putative Zn(II) 2Cys6 transcription factor (BC2). These 15 genes are specifically expressed during penetration into the host plant, defining an infection-specific gene cluster. A pORF3-GFP transcriptional fusion showed that the highly expressed ORF3 gene from the ACE1 cluster is only expressed in appressoria, as is ACE1. Phenotypic analysis of deletion or disruption mutants of SYN2 and RAP2 showed that they are not required for avirulence in Pi33 rice cultivars, unlike ACE1. Inactivation of other genes was unsuccessful because targeted gene replacement and disruption were inefficient at this locus. Overall, the ACE1 gene cluster displays an infection-specific expression pattern restricted to the penetration stage which is probably controlled at the transcriptional level and reflects regulatory networks specific to early stages of infection.
Early and specific gene expression triggered by rice resistance gene Pi33 in response to infection by ACE1 avirulent blast fungus E. Vergne, E. Ballini, S. Marques, B. Sidi Mammar, G. Droc, S. Gaillard, S. Bourot, R. DeRose, D. Tharreau, J.-L. Nottéghem, M.-H. Lebrun and J.-B. Morel
Summary Author for correspondence: J.-B. Morel Tel: +33 499624 837 Fax: +33 499624 822 Email: jbmorel@cirad.fr
Our view of genes involved in rice disease resistance is far from complete. Here we used a gene-for-gene relationship corresponding to the interaction between atypical avirulence gene ACE1 from Magnaporthe grisea and rice resistance gene Pi33 to better characterize early rice defence responses induced during such interaction.
New Phytologist (2007) 174: 159–171 doi : 10.1111/j.1469-8137.2007.01971.x
Rice genes differentially expressed during early stages of Pi33/ACE1 interaction were identified using DNA chip-based differential hybridization and QRT-PCR survey of the expression of known and putative regulators of disease resistance.
KEYWORDS: ACE1/Pi33 gene-for-gene interaction, defence response, Magnaporthe grisea, Oryza sativa (rice).
One hundred genes were identified as induced or repressed during rice defence response, 80% of which are novel, including resistance gene analogues. Pi33/ACE1 interaction also triggered the up-regulation of classical PR defence genes and a massive down-regulation of chlorophyll a/b binding genes. Most of these differentially expressed genes were induced or repressed earlier in Pi33/ACE1 interaction than in the gene–for–gene interaction involving Nipponbare resistant cultivar. Besides demonstrating that an ACE1/Pi33 interaction induced classical and specific expression patterns, this work provides a list of new genes likely to be involved in rice disease resistance.
The Lmgpi15 gene, encoding a component of the glycosylphosphatidylinositol anchor biosynthesis pathway, is required for morphogenesis and pathogenicity in Leptosphaeria maculans Estelle Remy, Michel Meyer, Françoise Blaise, Uwe K. Simon, Diana Kuhn, Mélanie Chabirand, Meritxell Riquelme, Marie-Hélène Balesdent and Thierry Rouxel
Summary Author for correspondence: Thierry Rouxel Tel: +33 1 30833229 Fax: +33 1 30833195 Email: rouxel@versailles.inra.fr
Random insertional mutagenesis was used to investigate pathogenicity determinants in Leptosphaeria maculans. One tagged nonpathogenic mutant, termed m20, was analysed in detail here.
The mutant phenotype was investigated by microscopic analyses of infected plant tissues and in vitro growth assays. Complementation and silencing experiments were used to identify the altered gene. Its function was determined by bioinformatics analyses, cell biology experiments and functional studies.
The mutant was blocked at the invasive growth phase after an unaffected initial penetration stage, and displayed a reduced growth rate and an aberrant hyphal morphology in vitro. The T-DNA insertion occurred in the intergenic region between two head-to-tail genes, leading to a complex deregulation of their expression. The unique gene accounting for the mutant phenotype was suggested to be the orthologue of the poorly conserved Saccharomyces cerevisiae gpi15, which encodes for one component of the glycosylphosphatidylinositol (GPI) anchor biosynthesis pathway. Consistent with this predicted function, a functional translational fusion with the green fluorescent protein (GFP) was targeted to the endoplasmic reticulum. Moreover, the mutant exhibited an altered cell wall and addition of glucosamine relieved growth defects.
It is concluded that the GPI anchor biosynthetic pathway is required for morphogenesis, cell wall integrity and pathogenicity in Leptosphaeria maculans.
New Phytologist (2008) 179: 1105–1120 doi: 10.1111/j.1469-8137.2008.02522.x
KEYWORDS: glycosylphosphatidylinositol (GPI) anchor biosynthesis, insertional mutagenesis, Leptosphaeria maculans, morphogenesis, oilseed rape, pathogenicity gene, stem canker.
Fusarium graminearum gene deletion mutants map1 and tri5 reveal similarities and differences in the pathogenicity requirements to cause disease on Arabidopsis and wheat floral tissue Alayne Cuzick, Martin Urban and Kim Hammond-Kosack
Summary Author for correspondence: Kim Hammond-Kosack Tel: +44 (0) 1582 763 133 ext 2240 Fax: +44 (0) 1582 760 981 Email: kim.hammondkosack@bbsrc.ac.uk
The Ascomycete pathogen Fusarium graminearum can infect all cereal species and lower grain yield, quality and safety. The fungus can also cause disease on Arabidopsis thaliana. In this study, the disease-causing ability of two F. graminearum mutants was analysed to further explore the parallels between the wheat (Triticum aestivum) and Arabidopsis floral pathosystems.
New Phytologist (2008) 177: 990–1000 doi: 10.1111/j.1469-8137.2007.02333.x
Wild-type F. graminearum (strain PH-1) and two isogenic transformants lacking either the mitogen-activated protein kinase MAP1 gene or the trichodiene synthase TRI5 gene were individually spray- or point-inoculated onto Arabidopsis and wheat floral tissue. Disease development was quantitatively assessed both macroscopically and microscopically and deoxynivalenol (DON) mycotoxin concentrations determined by enzyme-linked immunosorbent assay (ELISA).
Wild-type strain inoculations caused high levels of disease in both plant species and significant DON production. The map1 mutant caused minimal disease and DON accumulation in both hosts. The tri5 mutant, which is unable to produce DON, exhibited reduced pathogenicity on wheat ears, causing only discrete eye-shaped lesions on spikelets which failed to infect the rachis. By contrast, the tri5 mutant retained full pathogenicity on Arabidopsis floral tissue.
This study reveals that DON mycotoxin production is not required for F. graminearum to colonize Arabidopsis floral tissue.
KEYWORDS: Arabidopsis thaliana, deoxynivalenol, ear blight, Fusarium graminearum, Gibberella zeae, head scab, pathogenicity factor, Triticum aestivum. Gene ontology (GO) terms, GO:0009405 and GO:0044403.
Fusarium graminearum exploits ethylene signalling to colonize dicotyledonous and monocotyledonous plants X. Chen, A. Steed, S. Travella, B. Keller and P. Nicholson
Summary Author for correspondence: Paul Nicholson Tel: +44 (0) 1603 450 616 Email: paul.nicholson@bbsrc.ac.uk
Ethylene signalling affects the resistance of dicotyledonous plant species to diverse pathogens but almost nothing is known about the role of this pathway in monocotyledonous crop species. Fusarium graminearum causes Fusarium head blight (FHB) of cereals, contaminating grain with mycotoxins such as deoxynivalenol (DON). Very little is known about the mechanisms of resistance/susceptibility to this disease.
New Phytologist (2009) 182: 975–983 doi: 10.1111/j.1469-8137.2009.02821.x
Genetic and chemical genetic studies were used to examine the influence of ethylene (ET) signalling and perception on infection of dicotyledonous (Arabidopsis) andmonocotyledonous (wheat and barley) species by F. graminearum.
KEYWORDS: Arabidopsis, barley, deoxynivalenol (DON), ethylene, Fusarium head blight (FHB), Fusarium graminearum, wheat.
Arabidopsis mutants with reduced ET signalling or perception were more resistant to F. graminearum than wild-type, while mutants with enhanced ET production were more susceptible. These findings were confirmed by chemical genetic studies of Arabidopsis, wheat and barley. Attenuation of expression of EIN2 in wheat, a gene encoding a core component of ethylene signalling, reduced both disease symptoms and DON contamination of grain.
Fusarium graminearum appears to exploit ethylene signalling in both monocotyledonous and dicotyledonous species. This demonstration of translation from model to crop species provides a foundation for improving resistance of cereal crops to FHB through identification of allelic variation for components of the ethylenesignalling pathway.
Plant signalling components EDS1 and SGT1 enhance disease caused by the necrotrophic pathogen Botrytis cinerea Mohamed El Oirdi and Kamal Bouarab
Summary Author for correspondence: Kamal Bouarab Tel: +1 819 8217070 Fax: + 1 819 8218049 Email: Kamal.Bouarab@USherbrooke.ca New Phytologist (2007) 175 : 131–139 doi: 10.1111/j.1469-8137.2007.02086.x
KEYWORDS: antiapoptotic proteins, Botrytis cinerea, counter-defence strategies, EDS1, plant defences, SGT1, Solanaceae.
Botrytis cinerea is a necrotrophic fungus that causes grey mould on a wide range of food plants, especially grapevine, tomato, soft fruits and vegetables. This disease brings about important economic losses in both pre- and postharvest crops. Successful protection of host plants against this pathogen is severely hampered by a lack of resistance genes in the hosts and the considerable phenotypic diversity of the fungus.
The aim of this study was to test whether B. cinerea manipulates the immunitysignalling pathways in plants to restore its disease.
We showed that B. cinerea caused disease in Nicotiana benthamiana through the activation of two plant signalling genes, EDS1 and SGT1, which have been shown to be essential for resistance against biotrophic pathogens; and more interestingly, virus-induced gene silencing of these two plant signalling components enhanced N. benthamiana resistance to B. cinerea. Finally, plants expressing the baculovirus antiapoptotic protein p35 were more resistant to this necrotrophic pathogen than wild-type plants.
This work highlights a new strategy used by B. cinerea to establish disease. This information is important for the design of strategies to improve plant pathogen resistance.
Tracks for traffic: mirotubules in the plant pathogen Ustilago maydis Gero Steinberg
Summary Author for correspondence: Gero Steinberg Tel: +49 6421 178530 Fax: +49 6421 178599 Email: Gero.Steinberg@mpimarburg.mpg.de New Phytologist (2007) 174: 721–733 doi: 10.1111/j.1469-8137.2007.02072.x
KEYWORDS: dynein, hyphal growth, kinesin, microtubule array, organelle transport.
Pathogenic development of the corn smut fungus Ustilago maydis depends on the ability of the hypha to grow invasively. Extended hyphal growth and mitosis require microtubules, as revealed by recent studies on the microtubule cytoskeleton. Surprisingly, hyphal tip growth involves only two out of 10 kinesins. Kinesin-3 is responsible for tipdirected (anterograde) endosome motility of early endosomes, which are thought to support hyphal elongation by apical membrane recycling. In addition, kinesin-3, together with kinesin-1 and myosin-5, appear to deliver secretory vesicles to the hyphal tip. Kinesin-1 also affects endosome motility by targeting cytoplasmic dynein to microtubule plus ends. This plus-end localization of dynein is essential for cell body-directed (retrograde) endosome motility, but also allows force generation during spindle elongation in mitosis. Furthermore, kinesin-1 and dynein participate in the organization of the microtubule array, thereby building their own network of tracks for intracellular motility. The recent progress in understanding microtubule-based processes in U. maydis has revealed an unexpected complexity of motor functions essential for the virulence of this pathogen. Further studies on structural and regulatory requirements for motor activity should help identify novel targets for fungicide development.
Microbiological control of soil-borne phytopathogenic fungi with special emphasis on wilt-inducing Fusarium oxysporum Claude Alabouvette, Chantal Olivain, Quirico Migheli and Christian Steinberg
Summary Author for correspondence: Claude Alabouvette Tel: +33 380 693041 Email: ala@dijon.inra.fr
New Phytologist (2009) 184: 529–544 doi: 10.1111/j.1469-8137.2009.03014.x
KEYWORDS: biocontrol, biotrophy, competition, ecological fitness, induced resistance, plant defense reactions, priming, root colonization. root colonization.
Plant diseases induced by soil-borne plant pathogens are among the most difficult to control. In the absence of effective chemical control methods, there is renewed interest in biological control based on application of populations of antagonistic micro-organisms. In addition to Pseudomonas spp. and Trichoderma spp., which are the two most widely studied groups of biological control agents, the protective strains of Fusarium oxysporum represent an original model. These protective strains of F. oxysporum can be used to control wilt induced by pathogenic strains of the same species. Exploring the mechanisms involved in the protective capability of these strains is not only necessary for their development as commercial biocontrol agents but raises many basic questions related to the determinism of pathogenicity versus biocontrol capacity in the F. oxysporum species complex. In this paper, current knowledge regarding the interaction between the plant and the protective strains is reviewed in comparison with interactions between the plant and pathogenic strains. The success of biological control depends not only on plant–microbial interactions but also on the ecological fitness of the biological control agents.
The use of FLP-mediated recombination for the functional analysis of an effector gene family in the biotrophic smut fungus Ustilago maydis Yuliya Khrunyk, Karin Münch, Kerstin Schipper, Andrei N. Lupas and Regine Kahmann
Summary Author for correspondence: R. Kahmann Tel: +49 6421 178 501 Email: kahmann@mpimarburg.mpg.de
In the Ustilago maydis genome, several novel secreted effector proteins are encoded by gene families. Because of the limited number of selectable markers, the ability to carry out sequential gene deletions has limited the analysis of effector gene families that may have redundant functions.
Here, we established an inducible FLP-mediated recombination system in U. maydis that allows repeated rounds of gene deletion using a single selectable marker (HygR). To avoid genome rearrangements via FRT sites remaining in the genome after excision, different mutated FRT sites were introduced.
The FLP-mediated selectable marker-removal technique was successfully applied to delete a family of 11 effector genes (eff1) using five sequential rounds of recombination. We showed that expression of all 11 genes is up-regulated during the biotrophic phase. Strains carrying deletions of 9 or all 11 genes showed a significant reduction in virulence, and this phenotype could be partially complemented by the introduction of different members from the gene family, demonstrating redundancy.
The establishment of the FLP/FRT system in a plant pathogenic fungus paves the way for analyzing multigene families with redundant functions.
New Phytologist (2010) 187: 957–968 doi: 10.1111/j.1469-8137.2010.03413.x
Key words: FLP-mediated recombination, gene family, marker deletion, secreted effector, virulence.
Dynamic carbon transfer during pathogenesis of sunflower by the necrotrophic fungus Botrytis cinerea: from plant hexoses to mannitol Thierry Dulermo, Christine Rascle, Gaetan Chinnici, Elisabeth Gout, Richard Bligny and Pascale Cotton
Summary Author for correspondence: Pascale Cotton Tel: +33 472448030 Email: cotton@biomserv.univlyon1.fr
The main steps for carbon acquisition and conversion by Botrytis cinerea during pathogenesis of sunflower cotyledon were investigated here.
A sequential view of soluble carbon metabolites detected by NMR spectroscopy during infection is presented. Disappearance of plant hexoses and their conversion to fungal metabolites were investigated by expression analysis of an extended gene family of hexose transporters (Bchxts) and of the mannitol pathway, using quantitative PCR. In order to analyse the main fungal metabolic routes used by B. cinerea in real time, we performed, for the first time, in vivo NMR analyses during plant infection.
During infection, B. cinerea converts plant hexoses into mannitol. Expression analysis of the sugar porter gene family suggested predominance for transcription induced upon low glucose conditions and regulated according to the developmental phase. Allocation of plant hexoses by the pathogen revealed a conversion to mannitol, trehalose and glycogen for glucose and a preponderant transformation of fructose to mannitol by a more efficient metabolic pathway.
Uptake of plant hexoses by B. cinerea is based on a multigenic flexible hexose uptake system. Their conversion into mannitol, enabled by two simultaneously expressed pathways, generates a dynamic intracellular carbon pool.
New Phytologist (2009) 183: 1149–1162 doi: 10.1111/j.1469-8137.2009.02890.x
KEYWORDS: Botrytis, carbon metabolism, fungi, hexose transport gene family, in vivo NMR spectroscopy, mannitol pathway, necrotroph.
Expression profiling and functional analysis of Populus WRKY23 reveals a regulatory role in defense Valérie Levée, Ian Major, Caroline Levasseur, Laurence Tremblay, John MacKay and Armand Séguin
Summary Author for correspondence: Armand Séguin Tel: +1 418 648 5832 Email: armand.seguin@nrcan.gc.ca
New Phytologist (2009) 184: 48–70 doi: 10.1111/j.1469-8137.2009.02955.x
KEYWORDS: biotroph, forest tree, fungal pathogen, genomics, transcriptomic, WRKY.
WRKY transcription factors are key regulators that activate and fine-tune stress responses, including defense responses against pathogens. We isolated a poplar (Populus tremula×Populus alba) cDNA sequence, PtWRKY23, that encodes the ortholog of Arabidopsis WRKY23 and present the functional analysis of WRKY23, with emphasis on its potential role in resistance to rust infection.
To investigate the function of PtWRKY23, we examined PtWRKY23 expression after stress treatments by qRT-PCR and generated PtWRKY23-misexpressing plants. Transgenic plants were assessed for resistance to Melampsora rust and were analyzed using the poplar Affymetrix GeneChip® and histological techniques to study the consequences of PtWRKY23 misexpression.
PtWRKY23 is rapidly induced by Melampsora infection and elicitor treatments and poplars overexpressing and underexpressing PtWRKY23 were both more susceptible to Melampsora infection than wild type. Transcriptome analysis of PtWRKY23 overexpressors revealed a significant overlap with the Melampsora-infection response. Transcriptome analysis also suggests that PtWRKYI23 affects redox homeostasis and cell wall-related metabolism, which was confirmed by analyses that showed that PtWRKY23-misexpressing plants have altered peroxidase activity, apparent H2O2 accumulation and lignin deposition.
Our results show that PtWRKY23 affects resistance to MeIlampsora infection and that this may be caused by deregulation of genes that disrupt redox homeostasis and cell wall metabolism.
Natural genetic resources of Arabidopsis thaliana reveal a high prevalence and unexpected phenotypic plasticity of RPW8-mediated powdery mildew resistance Katharina Göllner, Patrick Schweizer, Yuling Bai and Ralph Panstruga
Summary Author for correspondence: Ralph Panstruga Tel: +49 (0)221 5062 316 Fax: +49 (0)221 5062 353 Email: panstrug@mpiz-koeln.mpg.de
Here, an approach based on natural genetic variation was adopted to analyse powdery mildew resistance in Arabidopsis thaliana.
Accessions resistant to multiple powdery mildew species were crossed with the susceptible Col-0 ecotype and inheritance of resistance was analysed. Histochemical staining was used to visualize archetypal plant defence responses such as callose deposition, hydrogen peroxide accumulation and host cell death in a subset of these ecotypes.
In six accessions, resistance was likely of polygenic origin while 10 accessions exhibited evidence for a single recessively or semi-dominantly inherited resistance locus. Resistance in the latter accessions was mainly manifested at the terminal stage of the fungal life cycle by a failure of abundant conidiophore production. The resistance locus of several of these ecotypes was mapped to a genomic region containing the previously analysed atypical RPW8 powdery mildew resistance genes. Gene silencing revealed that members of the RPW8 locus were responsible for resistance to Golovinomyces orontii in seven accessions.
These results suggest that broad-spectrum powdery mildew resistance in A. thaliana is predominantly of polygenic origin or based on RPW8 function. The findings shed new light on the natural variation of inheritance, phenotypic expression and pathogen range of RPW8-conditioned powdery mildew resistance.
New Phytologist (2008) 177: 725–742 doi: 10.1111/j.1469-8137.2007.02339.x
KEYWORDS: disease resistance, gene silencing, natural genetic variation, powdery mildew, RPW8
RPW8 and resistance to powdery mildew pathogens in natural populations of Arabidopsis lyrata T. H. Jorgensen and B. C. Emerson
Summary Author for correspondence: T. H. Jorgensen Tel: +44 (0) 1603 592269 Email: t.jorgensen@uea.ac.uk
It is not clear to what extent the orthologues of genes that are adaptively important in one species also contribute to adaptive variation in others. Here, we examine Arabidopsis lyrata to assess the functional and evolutionary significance of natural variation in an orthologue of the gene RPW8 known to be a major determinant of powdery mildew resistance in Arabidopsis thaliana.
New Phytologist (2009) 182: 984–993 doi: 10.1111/j.1469-8137.2009.02787.x
We assessed the sequence variation at RPW8 and the associated resistance reaction in populations of A. lyrata ssp. petraea. Neutrality tests were performed to understand the importance of local adaptation in maintaining variation at the locus.
KEYWORDS: Arabidopsis lyrata, Arabidopsis thaliana, Golovinomyces, orthologue, population structure, resistance, RPW8, selection.
Highly truncated RPW8 proteins were frequent in all populations and were associated with an increased risk of susceptibility. Haplotypes encoding full-length proteins were highly significantly associated with resistance. There were no signatures of selection at the species-wide level, but some evidence for positive selection in two populations.
RPW8 in A. lyrata appears to have a role in powdery mildew resistance, similar to its orthologue in A. thaliana. Unlike A. thaliana, A. lyrata contains a genetic component that can act independently of RPW8 to confer resistance to powdery mildew pathogens. Infrequent local selective sweeps may favour different alleles in different populations, and thereby contribute to the maintenance of species-wide variation at the locus.
Activation tagging of ADR2 conveys a spreading lesion phenotype and resistance to biotrophic pathogens Mourad A. M. Aboul-Soud, Xinwei Chen, Jeong-Gu Kang, Byung-Wook Yun, M. Usman Raja, Saad I. Malik and Gary J. Loake
Summary Author for correspondence: Gary J. Loake Tel: +44 0 131 650 5332 Email: gloake@ed.ac.uk
An Arabidopsis PR1::luciferase (LUC) transgenic line was transformed with activation T-DNA tags and the resulting population screened for dominant gain-of-function mutants exhibiting constitutive LUC activity.
LUC imaging identified activated disease resistance 2 (adr2), which exhibited slowly spreading lesions in the absence of pathogen challenge. Molecular, genetic and histochemical analysis was employed to characterize this mutant in detail.
adr2 plants constitutively expressed defence-related and antioxidant genes. Moreover, this line accrued increased quantities of salicylic acid (SA) and exhibited heightened mitogen-activated protein kinase activity. adr2 plants exhibited increased resistance against numerous biotrophic but not necrotrophic pathogens. The adr2 phenotype resulted from the overexpression of a Toll interleukin receptor (TIR) nucleotide binding site (NBS) leucine rich repeat (LRR) gene (At1g56510). Constitutive PR1 expression was completely abolished in adr2 nahG, adr2 npr1 and adr2 eds1 double mutants. Furthermore, heightened resistance against Hyaloperonospora arabidopsis Noco2 was compromised in adr2 nahG and adr2 eds1 double mutants but not in adr2 npr1, adr2 coi1 or adr2 etr1 plants.
These data imply that adr2-mediated resistance operates through an Enhanced Disease Susceptibility (EDS) and SA-dependent defence signalling network which functions independently from COI1 or ETR1.
New Phytologist (2009) 183: 1163–1175 doi: 10.1111/j.1469-8137.2009.02902.x
KEYWORDS: activation tagging, cell death, defence signalling, disease resistance, PR1 regulation, salicylic acid (SA).
Specificity and levels of nonhost resistance to nonadapted Blumeria graminis forms in barley Reza Aghnoum and Rients E. Niks
Summary Author for correspondence: Rients E. Niks Tel: +31 317 482 508 Email: rients.niks@wur.nl
The genetic basis of nonhost resistance of barley to nonadapted formae speciales of Blumeria graminis is not known, as there is no barley line that is susceptible to these nonadapted formae speciales, such as the wheat powdery mildew pathogen, Blumeria graminis f.sp. tritici (Bgt).
Barley accessions with rudimentary susceptibility to an isolate of the nonadapted Bgt were identified. Those accessions were intercrossed in two cycles and two lines, called SusBgtSC and SusBgtDC, with substantial susceptibility to Bgt at the seedling stage were selected.
The quantitative variation among barley accessions and in the progenies after convergent crossing suggests a polygenic basis for this nonhost resistance. Both lines allowed an unusually high level of haustorium formation and colony development by Bgt. The SusBgt lines and their ancestor lines also allowed haustorium formation and conidiation by four out of seven isolates of other nonadapted B. graminis forms. Analysis of the infection process suggested that nonhost resistance factors are specific to the form and developmental stage of B. graminis. Resistances to establishment (first haustorium), colonization (subsequent haustoria) and conidiation are not associated.
The lines developed will be of use in elucidating the genetic basis of nonhost resistance to Bgt in barley, and in gene expression and complementation studies on nonhost resistance.
New Phytologist (2010) 185: 275–284 doi: 10.1111/j.1469-8137.2009.03039.x
KEYWORDS: barley, Blumeria graminis, formae speciales, nonadapted pathogen, nonhost resistance, powdery mildew, pre-haustorial resistance.
Role of hydrogen peroxide during the interaction between the hemibiotrophic fungal pathogen Septoria tritici and wheat Nandini P. Shetty, Rahim Mehrabi, Henrik Lütken, Anna Haldrup, Gert H. J. Kema, David B. Collinge and Hans Jørgen Lyngs Jørgensen
Summary Author for correspondence: N. P. Shetty Tel: +45 35 28 23 23 Fax: +45 35 28 33 10 Email: nps@life.ku.dk
New Phytologist (2007) 174: 637 –647 doi: 10.1111/j.1469-8137.2007.02026.x
KEYWORDS: hemibiotrophic, hydrogen peroxide (H2O2), Mycosphaerella graminicola, Septoria tritici, stress, wheat (Triticum aestivum).
Hydrogen peroxide (H2O2) is reported to inhibit biotrophic but benefit necrotrophic pathogens. Infection by necrotrophs can result in a massive accumulation of H2O2 in hosts. Little is known of how pathogens with both growth types are affected (hemibiotrophs). The hemibiotroph, Septoria tritici, infecting wheat (Triticum aestivum) is inhibited by H2O2 during the biotrophic phase, but a large H2O2 accumulation occurs in the host during reproduction. Here, we infiltrated catalase, H2O2 or water into wheat during the biotrophic or the necrotrophic phase of S. tritici and studied the effect of infection on host physiology to get an understanding of the survival strategy of the pathogen. H2O2 removal by catalase at both early and late stages made plants more susceptible, whereas H2O2 made them more resistant. H2O2 is harmful to S. tritici throughout its life cycle, but it can be tolerated. The late accumulation of H2O2 is unlikely to result from down-regulation of photosynthesis, but probably originates from damage to the peroxisomes during the general tissue collapse, which is accompanied by release of soluble sugars in a susceptible cultivar.
De novo biosynthesis of defense root exudates in response to Fusarium attack in barley Arnaud Lanoue, Vincent Burlat, Gunnar J. Henkes, Imke Koch, Ulrich Schurr and Ursula S. R. Röse
Summary Author for correspondence: Arnaud Lanoue Tel: +33 (0) 247367214 Email: arnaud.lanoue@univ-tours.fr
New Phytologist (2010) 185: 577–588 doi: 10.1111/j.1469-8137.2009.03066.x
Despite recent advances in elucidation of natural products in root exudates, there are significant gaps in our understanding of the ecological significance of products in the rhizosphere.
Here, we investigated the potential of barley (Hordeum vulgare) to secrete defense root exudates when challenged by the soilborne pathogen Fusarium graminearum.
Liquid chromatography with photodiode array detection (LC-DAD) was used to profile induced small-molecular-weight exudates. Thus, t-cinnamic, p-coumaric, ferulic, syringic and vanillic acids were assigned to plant metabolism and were induced within 2 d after Fusarium inoculation. Biological tests demonstrated the ability of those induced root exudates to inhibit the germination of F. graminearum macroconidia. In vivo labeling experiments with 13CO2 revealed that the secreted t-cinnamic acid was synthesized de novo within 2 d of fungal infection. Simultaneously to its root exudation, t-cinnamic acid was accumulated in the roots. Microscopic analysis showed that nonlignin cell wall phenolics were induced not only in necrosed zones but in all root tissues.
Results suggest that barley plants under attack respond by de novo biosynthesis and secretion of compounds with antimicrobial functions that may mediate natural disease resistance.
KEYWORDS: barley (Hordeum vulgare), Fusarium graminearum, plant defense, plant–pathogen interaction, root exudates.
Action and reaction of host and pathogen during Fusarium head blight disease Stephanie Walter, Paul Nicholson and Fiona M. Doohan
Summary Author for correspondence: Stephanie Walter Tel: 0045 8999 3658 Email: stephanie.walter@agrsci.dk
New Phytologist (2010) 185: 54–66 doi: 10.1111/j.1469-8137.2009.03041.x
KEYWORDS: deoxynivalenol, disease resistance, Fusarium head blight (FHB), necrotroph, plant–pathogen interaction, trichothecene, wheat.
TheFusarium species Fusarium graminearum and Fusarium culmorum, which are responsible for Fusarium head blight (FHB) disease, reduce world-wide cereal crop yield and, as a consequence of their mycotoxin production in cereal grain, impact on both human and animal health. Their study is greatly promoted by the availability of the genomic sequence of F. graminearum and transcriptomic resources for both F. graminearum and its cereal hosts. Functional genomic, proteomic and metabolomic studies, in combination with targeted mutagenesis or transgenic studies, are unravelling the complex mechanisms involved in Fusarium infection, penetration and colonization of host tissues, and host avoidance thereof. This review illuminates and integrates emerging knowledge regarding the molecular crosstalk between Fusarium and its small-grain cereal hosts. An understanding of the complexity of the host–pathogen interactions will be instrumental in designing new efficient strategies for the control of FHB disease.
Arabidopsis thaliana plant defensin AtPDF1.1 is involved in the plant response to biotic stress Barbara M. A. De Coninck, Jan Sels, Esther Venmans, Wannes Thys, Inge J. W. M. Goderis, Delphine Carron, Stijn L. Delauré, Bruno P. A. Cammue, Miguel F. C. De Bolle and Janick Mathys
Summary Author for correspondence: Bruno Cammue Tel: +32 1632 9682 Email: bruno.cammue@biw.kuleuven.be
Previously, it was shown that the Arabidopsis thaliana plant defensins AtPDF1.1 (At1g75830) and AtPDF1.2a (At5g44420) exert in vitro antimicrobial properties and that their corresponding genes are expressed in seeds and induced in leaves upon pathogen attack, respectively.
In this study, the expression profile of both AtPDF1.1 and AtPDF1.2a is analysed in wild-type plants upon different stress-related treatments and the effect of modulation of their expression in transgenic plants is examined in both host and nonhost resistance.
AtPDF1.1, which was originally considered to be seed-specific, is demonstrated to be locally induced in leaves upon fungal attack and exhibits an expression profile distinct from that of AtPDF1.2a, a gene frequently used as marker for the ethylene/ jasmonate-mediated signaling pathway. Transgenic plants with modulated AtPDF1.1 or AtPDF1.2a gene expression show no altered phenotype upon Botrytis cinerea inoculation. However, constitutive overexpression of AtPDF1.1 in A. thaliana leads to a reduction in symptoms caused by the nonhost Cercospora beticola causing nonspreading spots on A. thaliana leaves.
These results indicate that AtPDF1.1 and AtPDF1.2a clearly differ regarding their expression profile and functionality in planta. It emphasizes the additional level of complexity and fine-tuning within the highly redundant plant defensin genes in A. thaliana.
New Phytologist (2010) 187: 1075–1088 doi: 10.1111/j.1469-8137.2010.03326.x
KEYWORDS: Arabidopsis thaliana, AtPDF1.1, AtPDF1.2a, Botrytis cinerea, Cercospora beticola, defense response, plant defensins.
Tête à tête inside a plant cell: establishing compatibility between plants and biotrophic fungi and oomycetes Richard J. O’Connell and Ralph Panstruga
Summary Author for correspondence: Ralph Panstruga Tel: +49-221-5062-316 Fax: +49-221-5062-353 Email: panstrug@mpiz-koeln.mpg.de
New Phytologist (2006) 171: 699–718 doi: 10.1111/j.1469-8137.2006.01829.x
KEYWORDS: Blumeria graminis, Colletotrichum, Hyaloperonospora parasitica, Magnaporthe grisea, Phytophthora, Uromyces fabae.
‘Compatibility’ describes the complementary relationship between a plant species and an adapted pathogen species that underlies susceptibility and which ultimately results in disease. Owing to elaborate surveillance systems and defence mechanisms on the plant side and a common lack of adaptation of many microbial pathogens, resistance is the rule and compatibility the exception for most plant–microbe combinations. While there has been major scientific interest in ‘resistance’ in the past decade, which has revealed many of its underlying molecular components, the analysis of ‘compatibility’, although intimately intertwined with ‘resistance’, has not been pursued with a similar intensity. Various recent studies, however, provide a first glimpse of the pivotal players and potential molecular mechanisms essential for compatibility in both the plant and parasite partners. In this review we highlight these findings with a particular emphasis on obligate biotrophic and hemibiotrophic fungal and oomycete pathogens and discuss novel strategies that might help to uncover further the molecular principles underlying compatibility to these highly specialized pathogens.
Frozen in time: a new method using cryo-scanning electron microscopy to visualize root–fungal interactions Steve Refshauge, Michelle Watt, Margaret E. McCully and Cheng X. Huang
Summary Author for correspondence: Michelle Watt Tel: +61 26246 4902 Fax: +61 26246 5399 Email: michelle.watt@csiro.au
New Phytologist (2006) 172: 369–374 doi: 10.1111/j.1469-8137.2006.01825.x
KEYWORDS: cryo-fixation, cryoplaning, cryo-scanning electron microscopy, deep etching, pathogen, Rhizoctonia solani, root–fungal interaction, root rot.
A new method of sample preparation for cryo-scanning electron microscopy was used to visualize internal infection of wheat (Triticum aestivum) roots by the pathogenic fungus Rhizoctonia solani AG-8. The new method retained fungal hyphae and root cells in situ in disintegrating root tissues, thus avoiding the distortions that can be introduced by conventional preparation by chemical fixation, dehydration and embedding.
Infected roots frozen in liquid nitrogen were cryo-planed and etched (sublimed) at −80°C for a critical length of time (up to 9 min) in the microscope column to reveal plant and fungal structures in three dimensions.
Root and fungal structures were well preserved irrespective of infection severity. Root and hyphal cell walls were clearly seen and hyphal architecture within and between root cells was preserved.
This rapid method permits three-dimensional in situ visualization of fungal invasion within roots and has broad application for examination of diseases caused by other necrotrophic fungi.
Multivesicular compartments proliferate in susceptible and resistant MLA12-barley leaves in response to infection by the biotrophic powdery mildew fungus Qianli An, Katrin Ehlers, Karl-Heinz Kogel, Aart J. E. Van Bel and Ralph Hückelhoven
Summary Author for correspondence: Ralph Hückelhoven Tel: +49 641 99 37494 Fax: +49 641 99 37499 Email: Ralph.Hueckelhoven@agrar.unigiessen.de
There is growing evidence that multivesicular bodies and cell wall-associated paramural bodies participate in the enhanced vesicle trafficking induced by pathogen attack.
Here, we performed transmission electron microscopy in combination with cytochemical localization of H2O2 to investigate multivesicular compartments during establishment of compatible interaction in susceptible barley (Hordeum vulgare) and during hypersensitive response in resistant MLA12-barley infected by the barley powdery mildew fungus (Blumeria graminis f. sp. hordei).
Multivesicular bodies, intravacuolar vesicle aggregates and paramural bodies proliferated in the penetrated epidermal cell during development of the fungal haustorium. These vesicular structures also proliferated at the periphery of intact cells, which were adjacent to the hypersensitive dying cells and deposited cell wall appositions associated with H2O2 accumulation. All plasmodesmata between intact cells and hypersensitive cells were constricted or blocked by cell wall appositions.
These results suggest that multivesicular compartments participate in secretion of building blocks for cell wall appositions not only to arrest fungal penetration but also to contain hypersensitive cell death through blocking plasmodesmata. They may also participate in internalization of damaged membranes, deleterious materials, nutrients, elicitors and elicitor receptors.
New Phytologist (2006) 172: 563–576 doi: 10.1111/j.1469-8137.2006.01844.x
KEYWORDS: endocytosis, exocytosis, hypersensitive response, multivesicular body, paramural body, plasmodesmata, powdery mildew, programmed cell death.
Phytotoxic Nep1-like proteins from the necrotrophic fungus Botrytis cinerea associate with membranes and the nucleus of plant cells Alexander Schouten, Peter Van Baarlen and Jan A. L. Van Kan
Summary Author for correspondence: Jan A. L. van Kan Tel: +31 317483126 Fax: +31 317483412 Email: jan.vankan@wur.nl
Nep1-like proteins (NLPs), produced by an array of unrelated microorganisms, are phytotoxic for dicotyledonous plant cells but their mode of action has not yet been established.
Two paralogous NLPs from the necrotrophic plant pathogenic fungus Botrytis cinerea were characterized, designated BcNEP1 and BcNEP2. Both proteins were produced in the heterologous host Pichia pastoris and purified to homogeneity. The localization of fluorescently labelled proteins was studied and mechanisms of cell death were investigated in protoplasts and suspension cells.
Purified BcNEP1 and BcNEP2 caused necrosis in all dicotyledonous plant species tested, but not in monocotyledons. A synthetic heptapeptide comprising a sequence (GHRHDWE) that is conserved in all NLPs did not cause symptoms and was unable to interfere with necrosis induction by BcNEP1 and BcNEP2 proteins. Fluorescently labelled BcNEP1 and BcNEP2 proteins were associated with plasma membranes and the nuclear envelope, as well as in the nucleolus of responding plant cells. A strong hydrogen peroxide (H2O2) accumulation was observed in chloroplasts. The death process was characterized by TUNEL assays as apoptosis, necrosis or intermediate forms of both. BcNEP1- and BcNEP2-induced cell death execution could not be abolished by specific inhibitors.
These results provide further information on mechanisms of NLP-inflicted cell death.
New Phytologist (2008) 177: 493–505 doi: 10.1111/j.1469-8137.2007.02274.x
KEYWORDS: grey mould, heterologous expression, necrosis, necrotroph, programmed cell death (PCD).
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