Large-Scale Phenomics Identifies Primary and Fine-Tuning Roles for CRKs in Responses Related to Oxidative Stress.
Bourdais, G. ; Burdiak, P. ; Gauthier, A. ; Nitsch, L.M.C. ; Salojärvi, J. ; Rayapuram, C. ; Idänheimo, N. ; Hunter, K. ; Kimura, S. ; Merilo, E. ; Vaattovaara, A. ; Oracz, K. ; Kaufholdt, D. ; Pallon, A. ; Anggoro, D.T. ; Glów, D. ; Lowe, J. ; Zhou, J. ; Mohammadi, O. ; Puukko, T. ; Albert, A. ; Lang, H. ; Ernst, D. ; Kollist, H. ; Brosché, M. ; Durner, J. ; Borst, J.W. ; Collinge, D.B. ; Karpinski, S. ; Lyngkjær, M.F. ; Robatzek, S. ; Wrzaczek, M. ; Kangasjärvi, J. - \ 2015
Plos Genetics 11 (2015)7. - ISSN 1553-7404
receptor-like kinase - multiple sequence alignment - arabidopsis-thaliana - cell-death - protein-kinase - transcriptional regulation - pseudomonas-syringae - flagellin perception - light acclimation - stomatal immunity
Cysteine-rich receptor-like kinases (CRKs) are transmembrane proteins characterized by the presence of two domains of unknown function 26 (DUF26) in their ectodomain. The CRKs form one of the largest groups of receptor-like protein kinases in plants, but their biological functions have so far remained largely uncharacterized. We conducted a large-scale phenotyping approach of a nearly complete crk T-DNA insertion line collection showing that CRKs control important aspects of plant development and stress adaptation in response to biotic and abiotic stimuli in a non-redundant fashion. In particular, the analysis of reactive oxygen species (ROS)-related stress responses, such as regulation of the stomatal aperture, suggests that CRKs participate in ROS/redox signalling and sensing. CRKs play general and fine-tuning roles in the regulation of stomatal closure induced by microbial and abiotic cues. Despite their great number and high similarity, large-scale phenotyping identified specific functions in diverse processes for many CRKs and indicated that CRK2 and CRK5 play predominant roles in growth regulation and stress adaptation, respectively. As a whole, the CRKs contribute to specificity in ROS signalling. Individual CRKs control distinct responses in an antagonistic fashion suggesting future potential for using CRKs in genetic approaches to improve plant performance and stress tolerance.
Signal Transduction by a Fungal NOD-Like Receptor Based on Propagation of a Prion Amyloid Fold
Daskalov, A. ; Habenstein, B. ; Martinez, D. ; Debets, A.J.M. ; Sabate, R. ; Loquet, A. ; Saupe, S.J. - \ 2015
PloS Biology 13 (2015)2. - ISSN 1545-7885
het-s prion - state nmr-spectroscopy - non-self recognition - heterokaryon incompatibility gene - glycolipid transfer protein - podospora-anserina - vegetative incompatibility - cell-death - het-s(218-289) prion - diversity
In the fungus Podospora anserina, the [Het-s] prion induces programmed cell death by activating the HET-S pore-forming protein. The HET-s ß-solenoid prion fold serves as a template for converting the HET-S prion-forming domain into the same fold. This conversion, in turn, activates the HET-S pore-forming domain. The gene immediately adjacent to het-S encodes NWD2, a Nod-like receptor (NLR) with an N-terminal motif similar to the elementary repeat unit of the ß-solenoid fold. NLRs are immune receptors controlling cell death and host defense processes in animals, plants and fungi. We have proposed that, analogously to [Het-s], NWD2 can activate the HET-S pore-forming protein by converting its prion-forming region into the ß-solenoid fold. Here, we analyze the ability of NWD2 to induce formation of the ß-solenoid prion fold. We show that artificial NWD2 variants induce formation of the [Het-s] prion, specifically in presence of their cognate ligands. The N-terminal motif is responsible for this prion induction, and mutations predicted to affect the ß-solenoid fold abolish templating activity. In vitro, the N-terminal motif assembles into infectious prion amyloids that display a structure resembling the ß-solenoid fold. In vivo, the assembled form of the NWD2 N-terminal region activates the HET-S pore-forming protein. This study documenting the role of the ß-solenoid fold in fungal NLR function further highlights the general importance of amyloid and prion-like signaling in immunity-related cell fate pathways
Sequestration of fatty acids in triglycerides prevents endoplasmic reticulum stress in an in vitro model of cardiomyocyte lipotoxicity
Bosma, M. ; Dapito, D.H. ; Drosatos-Tampakaki, Z. ; Huiping-Son, N. ; Huang, L.S. ; Kersten, A.H. ; Drosatos, K. ; Goldberg, I.J. - \ 2014
Biochimica et Biophysica Acta. Molecular and Cell Biology of Lipids 1841 (2014)12. - ISSN 1388-1981 - p. 1648 - 1655.
hormone-sensitive lipase - activated receptor-gamma - skeletal-muscle - insulin-resistance - cardiac myocytes - transgenic mice - cell-death - lipid-metabolism - heart-failure - ppar-alpha
We used human cardiomyocyte-derived cells to create an in vitro model to study lipid metabolism and explored the effects of PPAR gamma, ACSL1 and ATGL on fatty acid-induced ER stress. Compared to oleate, palmitate treatment resulted in less intracellular accumulation of lipid droplets and more ER stress, as measured by upregulation of CHOP, ATF6 and GRP78 gene expression and phosphorylation of eukaryotic initiation factor 2a (ElF2a). Both ACSL1 and PPAR gamma adenovirus-mediated expression augmented neutral lipid accumulation and reduced palmitate-induced upregulation of ER stress markers to levels similar to those in the oleate and control treatment groups. This suggests that increased channeling of non-esterffied free fatty acids (NEFA) towards storage in the form of neutral lipids in lipid droplets protects against palmitate-induced ER stress. Overexpression of ATGL in cells incubated with oleate-containing medium increased NEFA release and stimulated expression of ER stress markers. Thus, inefficient creation of lipid droplets as well greater release of stored lipids induces ER stress. (C) 2014 Elsevier B.V. All rights reserved.
Natural Variation of Heterokaryon Incompatibility Gene het-c in Podospora anserina Reveals Diversifying Selection
Bastiaans, E. ; Debets, A.J.M. ; Aanen, D.K. ; Diepeningen, A.D. van; Saupe, S.J. ; Paoletti, M. - \ 2014
Molecular Biology and Evolution 31 (2014)4. - ISSN 0737-4038 - p. 962 - 974.
plant immune-system - glycolipid transfer protein - chestnut blight fungus - amino-acid sites - vegetative incompatibility - neurospora-crassa - cell-death - cryphonectria-parasitica - membrane interaction - filamentous fungi
In filamentous fungi, allorecognition takes the form of heterokaryon incompatibility, a cell death reaction triggered when genetically distinct hyphae fuse. Heterokaryon incompatibility is controlled by specific loci termed het-loci. In this article, we analyzed the natural variation in one such fungal allorecognition determinant, the het-c heterokaryon incompatibility locus of the filamentous ascomycete Podospora anserina. The het-c locus determines an allogenic incompatibility reaction together with two unlinked loci termed het-d and het-e. Each het-c allele is incompatible with a specific subset of the het-d and het-e alleles. We analyzed variability at the het-c locus in a population of 110 individuals, and in additional isolates from various localities. We identified a total of 11 het-c alleles, which define 7 distinct incompatibility specificity classes in combination with the known het-d and het-e alleles. We found that the het-c allorecognition gene of P. anserina is under diversifying selection. We find a highly unequal allele distribution of het-c in the population, which contrasts with the more balanced distribution of functional groups of het-c based on their allorecognition function. One explanation for the observed het-c diversity in the population is its function in allorecognition. However, alleles that are most efficient in allorecognition are rare. An alternative and not exclusive explanation for the observed diversity is that het-c is involved in pathogen recognition. In Arabidopsis thaliana, a homolog of het-c is a pathogen effector target, supporting this hypothesis. We hypothesize that the het-c diversity in P. anserina results from both its functions in pathogen-defense, and allorecognition
Activation tagging of ATHB13 in Arabidopsis thaliana confers broad-spectrum disease resistance
Gao, D. ; Huibers, R.P. ; Chen, X. ; Loonen, A.E.H.M. ; Visser, R.G.F. ; Wolters, A.M.A. ; Bai, Y. - \ 2014
Plant Molecular Biology 86 (2014)6. - ISSN 0167-4412 - p. 641 - 653.
vegetative storage protein - powdery mildew resistance - transcription factors hahb1 - nudix hydrolase - salicylic-acid - plant defense - cell-death - hd-zip - oidium-neolycopersici - expression patterns
Powdery mildew species Oidium neolycopersici (On) can cause serious yield losses in tomato production worldwide. Besides on tomato, On is able to grow and reproduce on Arabidopsis. In this study we screened a collection of activation-tagged Arabidopsis mutants and identified one mutant, 3221, which displayed resistance to On, and in addition showed a reduced stature and serrated leaves. Additional disease tests demonstrated that the 3221 mutant exhibited resistance to downy mildew (Hyaloperonospora arabidopsidis) and green peach aphid (Myzus persicae), but retained susceptibility to bacterial pathogen Pseudomonas syringae pv tomato DC3000. The resistance trait and morphological alteration were mutually linked in 3221. Identification of the activation tag insertion site and microarray analysis revealed that ATHB13, a homeodomain-leucine zipper (HD-Zip) transcription factor, was constitutively overexpressed in 3221. Silencing of ATHB13 in 3221 resulted in the loss of both the morphological alteration and resistance, whereas overexpression of the cloned ATHB13 in Col-0 and Col-eds1-2 backgrounds resulted in morphological alteration and resistance. Microarray analysis further revealed that overexpression of ATHB13 influenced the expression of a large number of genes. Previously, it was reported that ATHB13-overexpressing lines conferred tolerance to abiotic stress. Together with our results, it appears that ATHB13 is involved in the crosstalk between abiotic and biotic stress resistance pathways.
Analysis of Tomato spotted wilt virus NSs protein indicates the importance of the N-terminal for avirulence and RNA silencing suppression
Ronde, D. de; Pasquier, A. ; Ying, S. ; Butterbach, P.B.E. ; Lohuis, D. ; Kormelink, R.J.M. - \ 2014
Molecular Plant Pathology 15 (2014)2. - ISSN 1464-6722 - p. 185 - 195.
turnip-crinkle-virus - cell-death - hypersensitive response - nonstructural protein - coat protein - tsw gene - resistance - arabidopsis - expression - plants
Recently, Tomato spotted wilt virus (TSWV) nonstructural protein NSs has been identified unambiguously as an avirulence (Avr) determinant for Tomato spotted wilt (Tsw)-based resistance. The observation that NSs from two natural resistance-breaking isolates had lost RNA silencing suppressor (RSS) activity and Avr suggested a link between the two functions. To test this, a large set of NSs mutants was generated by alanine substitutions in NSs from resistance-inducing wild-type strains (NSsRI), amino acid reversions in NSs from resistance-breaking strains (NSsRB), domain deletions and swapping. Testing these mutants for their ability to suppress green fluorescent protein (GFP) silencing and to trigger a Tsw-mediated hypersensitive response (HR) revealed that the two functions can be separated. Changes in the N-terminal domain were found to be detrimental for both activities and indicated the importance of this domain, additionally supported by domain swapping between NSsRI and NSsRB. Swapping domains between the closely related Tospovirus Groundnut ringspot virus (GRSV) NSs and TSWV NSsRI showed that Avr functionality could not simply be transferred between species. Although deletion of the C-terminal domain rendered NSs completely dysfunctional, only a few single-amino-acid mutations in the C-terminus affected both functions. Mutation of a GW/WG motif (position 17/18) rendered NSs completely dysfunctional for RSS and Avr activity, and indicated a putative interaction between NSs and Argonaute 1 (AGO1), and its importance in TSWV virulence and viral counter defence against RNA interference.
Identification of candidate genes required for susceptibility to powdery or downy mildew in cucumber
Schouten, H.J. ; Krauskopf, J. ; Visser, R.G.F. ; Bai, Y. - \ 2014
Euphytica 200 (2014)3. - ISSN 0014-2336 - p. 475 - 486.
quantitative trait loci - sativus l. - disease resistance - mlo-gene - qtl analysis - cell-death - arabidopsis - defense - protein - barley
Powdery mildew (PM, caused by Podosphaera fusca) and downy mildew (DM, caused by Pseudoperonospora cubensis) are important diseases of cucumber (Cucumis sativus). Breeding for resistance has been undertaken since the 1940s, but underlying resistance genes have not been functionally analysed yet. The published genome sequence of cucumber catalyses the search for such genes. Genetic studies have indicated that resistances to PM and DM in cucumber are often inherited recessively, which indicates the presence of susceptibility genes (S-genes). Therefore we analyzed the cucumber genome for homologs of functionally proven S-genes known from other plant species. We identified 13 MLO-like genes in cucumber, three of which cluster in Clade V, the clade that contains all known MLO-like susceptibility genes to powdery mildews in other dicots. The expression of one of these three genes, CsaMLO1, located on chromosome 1, was upregulated after PM inoculation. It co-localizes with a QTL for PM resistance previously identified. Also homologs of the susceptibility genes PMR4 and PMR5 are located at this QTL. The second MLO-like gene from Clade V (CsaMLO8) resides in a recessively inherited major QTL for PM resistance at the bottom of chromosome 5, together with a PMR6-like gene. Two major QTL for DM recessive resistance at the top of chromosome 5 co-localize with CsaDMR6-2, which is homologous to the DMR6 susceptibility gene in Arabidopsis. This study has identified several candidate genes for susceptibility to PM and DM in cucumber that may explain QTL for recessively inherited resistance, reported earlier.
Does autophagy mediate age-dependent effect of dietary restriction responses in the filamentous fungus Podospora anserina?
Diepeningen, A.D. van; Engelmoer, D.J.P. ; Sellem, C.H. ; Huberts, D.H.E.W. ; Slakhorst, S.M. ; Sainsard-Chanet, A. ; Zwaan, B.J. ; Hoekstra, R.F. ; Debets, A.J.M. - \ 2014
Philosophical Transactions of the Royal Society B. Biological sciences 369 (2014)1646. - ISSN 0962-8436
life-span extension - calorie restriction - cell-death - saccharomyces-cerevisiae - mitochondrial fission - uth1 gene - senescence - plasmid - yeast - degradation
Autophagy is a well-conserved catabolic process, involving the degradation of a cell's own components through the lysosomal/vacuolar machinery. Autophagy is typically induced by nutrient starvation and has a role in nutrient recycling, cellular differentiation, degradation and programmed cell death. Another common response in eukaryotes is the extension of lifespan through dietary restriction (DR). We studied a link between DR and autophagy in the filamentous fungus Podospora anserina, a multicellular model organism for ageing studies and mitochondrial deterioration. While both carbon and nitrogen restriction extends lifespan in P. anserina, the size of the effect varied with the amount and type of restricted nutrient. Natural genetic variation for the DR response exists. Whereas a switch to carbon restriction up to halfway through the lifetime resulted in extreme lifespan extension for wild-type P. anserina, all autophagy-deficient strains had a shorter time window in which ageing could be delayed by DR. Under nitrogen limitation, only PaAtg1 and PaAtg8 mediate the effect of lifespan extension; the other autophagy-deficient mutants PaPspA and PaUth1 had a similar response as wild-type. Our results thus show that the ageing process impinges on the DR response and that this at least in part involves the genetic regulation of autophagy
The leucine-rich repeat receptor kinase BIR2 is a negative regulator of BAK1 in plant immunity
Halter, T. ; Imkampe, J. ; Mazzotta, S. ; Wierzba, M. ; Postel, S. ; Bücherl, C.A. ; Kiefer, C. ; Stahl, M. ; Chinchilla, D. ; Wang, X. ; Nürnberger, T. ; Zipfel, C. ; Clouse, S. ; Borst, J.W. ; Boeren, S. ; Vries, S.C. de; Tax, F. ; Kemmerling, B. - \ 2014
Current Biology 24 (2014)2. - ISSN 0960-9822 - p. 134 - 143.
innate immunity - cell-death - necrotrophic pathogens - molecular-patterns - protein-kinase - arabidopsis - complex - bri1 - perception - activation
Background Transmembrane leucine-rich repeat (LRR) receptors are commonly used innate immune receptors in plants and animals but can also sense endogenous signals to regulate development. BAK1 is a plant LRR-receptor-like kinase (RLK) that interacts with several ligand-binding LRR-RLKs to positively regulate their functions. BAK1 is involved in brassinosteroid-dependent growth and development, innate immunity, and cell-death control by interacting with the brassinosteroid receptor BRI1, immune receptors, such as FLS2 and EFR, and the small receptor kinase BIR1, respectively. Results Identification of in vivo BAK1 complex partners by LC/ESI-MS/MS uncovered two novel BAK1-interacting RLKs, BIR2 and BIR3. Phosphorylation studies revealed that BIR2 is unidirectionally phosphorylated by BAK1 and that the interaction between BAK1 and BIR2 is kinase-activity dependent. Functional analyses of bir2 mutants show differential impact on BAK1-regulated processes, such as hyperresponsiveness to pathogen-associated molecular patterns (PAMP), enhanced cell death, and resistance to bacterial pathogens, but have no effect on brassinosteroid-regulated growth. BIR2 interacts constitutively with BAK1, thereby preventing interaction with the ligand-binding LRR-RLK FLS2. PAMP perception leads to BIR2 release from the BAK1 complex and enables the recruitment of BAK1 into the FLS2 complex. Conclusions Our results provide evidence for a new regulatory mechanism for innate immune receptors with BIR2 acting as a negative regulator of PAMP-triggered immunity by limiting BAK1-receptor complex formation in the absence of ligands.
Two for all: receptor-associated kinases SOBIR1 and BAK1
Liebrand, T.W.H. ; Burg, H.A. van den; Joosten, M.H.A.J. - \ 2014
Trends in Plant Science 19 (2014)2. - ISSN 1360-1385 - p. 123 - 132.
plant innate immunity - pattern-recognition receptors - ethylene-inducing xylanase - arabidopsis-thaliana - cladosporium-fulvum - defense responses - cell-death - signaling pathways - plasma-membrane - protein-kinase
Leucine-rich repeat-receptor-like proteins (LRR-RLPs) are ubiquitous cell surface receptors lacking a cytoplasmic signalling domain. For most of these LRR-RLPs, it remained enigmatic how they activate cellular responses upon ligand perception. Recently, the LRR-receptor-like kinase (LRR-RLK) SUPPRESSOR OF BIR1-1 (SOBIR1) was shown to be essential for triggering defence responses by certain LRR-RLPs that act as immune receptors. In addition to SOBIR1, the regulatory LRR-RLK BRI1-ASSOCIATED KINASE-1 (BAK1) is also required for LRR-RLP function. Here, we compare the roles of SOBIR1 and BAK1 as regulatory LRR-RLKs in immunity and development. BAK1 has a general regulatory role in plasma membrane-associated receptor complexes comprising LRR-RLPs and/or LRR-RLKs. By contrast, SOBIR1 appears to be specifically required for the function of receptor complexes containing LRR-RLPs.
Phellinus linteus polysaccharide extracts increase the mitochondrial membrane potential and cause apoptotic death of THP-1 monocytes
Griensven, L.J.L.D. van; Verhoeven, H.A. - \ 2013
Chinese Medicine 8 (2013). - ISSN 1749-8546 - 13 p.
trail-induced apoptosis - cell-death - prostate-cancer - beta-glucans - tumor-growth - macrophage - necrosis - protein - fungi - differentiation
Background: The differentiation resp. death of human monocytic THP-1 cells induced by polysaccharide extracts of the medicinal mushrooms Phellinus linteus, Agaricus bisporus and Agaricus brasiliensis have been studied. This study aims to identify leads for the causal effects of these mushroom components on cell differentiation and death. Methods: THP-1 cells were treated with different polysaccharide extracts of mushrooms and controls. Morphological effects were observed by light microscopy. Flow cytometry was applied to follow the cell differentiation by cell cycle shifts after staining with propidium iodide, changes of mitochondrial membrane potential after incubation with JC-1, and occurrence of intracellular reactive oxygen species after incubation with hydroethidine. Principal component analysis of the data was performed to evaluate the cellular effects of the different treatments. Results: P. linteus polysaccharide extracts induced dose-dependent apoptosis of THP-1 cells within 24 h, while A. bisporus and A. brasiliensis polysaccharide extracts caused differentiation into macrophages. A pure P. linteus polysaccharide had no effect. Apoptosis was inhibited by preincubating THP-1 cells with human serum. The principal component analysis revealed that P. linteus, A. bisporus and A. brasiliensis polysaccharide extracts increased reactive oxygen species production. Both A. bisporus and A. brasiliensis polysaccharide extracts decreased the mitochondrial membrane potential, while this was increased by P. linteus polysaccharide extracts. Conclusions: P. linteus polysaccharide extracts caused apoptosis of THP-1 monocytes while A. bisporus and A. brasiliensis polysaccharide extracts caused these cells to differentiate into macrophages. The protective effects of human serum suggested that P. linteus polysaccharide extract induced apoptosis by extrinsic pathway, i.e. by binding to the TRAIL receptor. The mitochondrial membrane potential together with reactive oxygen species seems to play an important role in cell differentiation and cell death.
Stepwise artificial evolution of a plant disease resistance gene
Harris, J.H. ; Slootweg, E.J. ; Goverse, A. ; Baulcombe, D.C. - \ 2013
Proceedings of the National Academy of Sciences of the United States of America 110 (2013)52. - ISSN 0027-8424 - p. 21189 - 21194.
nb-lrr protein - tobacco-mosaic-virus - rich repeat domains - cell-death - arabidopsis-thaliana - immune receptors - coiled-coil - hypersensitive response - mediated resistance - pathogen effectors
Genes encoding plant nucleotide-binding leucine-rich repeat (NB-LRR) proteins confer dominant resistance to diverse pathogens. The wild-type potato NB-LRR protein Rx confers resistance against a single strain of potato virus X (PVX), whereas LRR mutants protect against both a second PVX strain and the distantly related poplar mosaic virus (PopMV). In one of the Rx mutants there was a cost to the broad-spectrum resistance because the response to PopMV was transformed from a mild disease on plants carrying wild-type Rx to a trailing necrosis that killed the plant. To explore the use of secondary mutagenesis to eliminate this cost of broad-spectrum resistance, we performed random mutagenesis of the N-terminal domains of this broad-recognition version of Rx and isolated four mutants with a stronger response against the PopMV coat protein due to enhanced activation sensitivity. These mutations are located close to the nucleotide-binding pocket, a highly conserved structure that likely controls the “switch” between active and inactive NB-LRR conformations. Stable transgenic plants expressing one of these versions of Rx are resistant to the strains of PVX and the PopMV that previously caused trailing necrosis. We conclude from this work that artificial evolution of NB-LRR disease resistance genes in crops can be enhanced by modification of both activation and recognition phases, to both accentuate the positive and eliminate the negative aspects of disease resistance.
Ve1-mediated resistance against Verticillium does not involve a hypersensitive response in Arabidopsis
Zhang, Z. ; Esse, H.P. van; Damme, M. van; Fradin, E.F. ; Liu, Chun-Ming ; Thomma, B.P.H.J. - \ 2013
Molecular Plant Pathology 14 (2013)7. - ISSN 1464-6722 - p. 719 - 727.
ethylene-inducing xylanase - receptor-like proteins - gated ion-channel - disease resistance - rhynchosporium-secalis - functional-analysis - defense responses - gene family - tomato ve1 - cell-death
The recognition of pathogen effectors by plant immune receptors leads to the activation of immune responses that often include a hypersensitive response (HR): rapid and localized host cell death surrounding the site of attempted pathogen ingress. We have demonstrated previously that the recognition of the Verticillium dahliae effector protein Ave1 by the tomato immune receptor Ve1 triggers an HR in tomato and tobacco. Furthermore, we have demonstrated that tomato Ve1 provides Verticillium resistance in Arabidopsis upon Ave1 recognition. In this study, we investigated whether the co-expression of Ve1 and Ave1 in Arabidopsis results in an HR, which could facilitate a forward genetics screen. Surprisingly, we found that the co-expression of Ve1 and Ave1 does not induce an HR in Arabidopsis. These results suggest that an HR may occur as a consequence of Ve1/Ave1-induced immune signalling in tomato and tobacco, but is not absolutely required for Verticillium resistance.
Loss of Function in Mlo Orthologs Reduces Susceptibility of Pepper and Tomato to Powdery Mildew Disease Caused by Leveillula taurica
Zheng, Z. ; Nonomura, T. ; Appiano, M. ; Pavan, S.N.C. ; Matsuda, Y. ; Toyoda, H. ; Wolters, A.M.A. ; Visser, R.G.F. ; Bai, Y. - \ 2013
PLoS ONE 8 (2013)7. - ISSN 1932-6203 - 14 p.
real-time pcr - capsicum-annuum - subcellular-localization - nonhost resistance - gene-expression - oryza-sativa - cell-death - barley - family - identification
Powdery mildew disease caused by Leveillula taurica is a serious fungal threat to greenhouse tomato and pepper production. In contrast to most powdery mildew species which are epiphytic, L. taurica is an endophytic fungus colonizing the mesophyll tissues of the leaf. In barley, Arabidopsis, tomato and pea, the correct functioning of specific homologues of the plant Mlo gene family has been found to be required for pathogenesis of epiphytic powdery mildew fungi. The aim of this study was to investigate the involvement of the Mlo genes in susceptibility to the endophytic fungus L. taurica. In tomato (Solanum lycopersicum), a loss-of-function mutation in the SlMlo1 gene results in resistance to powdery mildew disease caused by Oidium neolycopersici. When the tomato Slmlo1 mutant was inoculated with L. taurica in this study, it proved to be less susceptible compared to the control, S. lycopersicum cv. Moneymaker. Further, overexpression of SlMlo1 in the tomato Slmlo1 mutant enhanced susceptibility to L. taurica. In pepper, the CaMlo2 gene was isolated by applying a homology-based cloning approach. Compared to the previously identified CaMlo1 gene, the CaMlo2 gene is more similar to SlMlo1 as shown by phylogenetic analysis, and the expression of CaMlo2 is up-regulated at an earlier time point upon L. taurica infection. However, results of virus-induced gene silencing suggest that both CaMlo1 and CaMlo2 may be involved in the susceptibility of pepper to L. taurica. The fact that overexpression of CaMlo2 restored the susceptibility of the tomato Slmlo1 mutant to O. neolycopersici and increased its susceptibility to L. taurica confirmed the role of CaMlo2 acting as a susceptibility factor to different powdery mildews, though the role of CaMlo1 as a co-factor for susceptibility cannot be excluded.
Receptor-like kinase SOBIR1/EVR interacts with receptor-like proteins in plant immunity against fungal infection
Liebrand, T.W.H. ; Berg, G.C.M. van den; Zhang, Z. ; Smit, P. ; Cordewener, J.H.G. ; America, A.H.P. ; Sklenar, J. ; Jones, A.M.E. ; Tameling, W.I.L. ; Robatzek, S. ; Thomma, B.P.H.J. ; Joosten, M.H.A.J. - \ 2013
Proceedings of the National Academy of Sciences of the United States of America 110 (2013)24. - ISSN 0027-8424 - p. 10010 - 10015.
innate immunity - cladosporium-fulvum - verticillium resistance - citrx thioredoxin - plasma-membrane - tomato ve1 - cell-death - arabidopsis - perception - flagellin
The plant immune system is activated by microbial patterns that are detected as nonself molecules. Such patterns are recognized by immune receptors that are cytoplasmic or localized at the plasma membrane. Cell surface receptors are represented by receptor-like kinases (RLKs) that frequently contain extracellular leucine-rich repeats and an intracellular kinase domain for activation of downstream signaling, as well as receptor-like proteins (RLPs) that lack this signaling domain. It is therefore hypothesized that RLKs are required for RLPs to activate downstream signaling. The RLPs Cf-4 and Ve1 of tomato (Solanum lycopersicum) mediate resistance to the fungal pathogens Cladosporium fulvum and Verticillium dahliae, respectively. Despite their importance, the mechanism by which these immune receptors mediate downstream signaling upon recognition of their matching ligand, Avr4 and Ave1, remained enigmatic. Here we show that the tomato ortholog of the Arabidopsis thaliana RLK Suppressor Of BIR1-1/Evershed (SOBIR1/EVR) and its close homolog S. lycopersicum (Sl)SOBIR1-like interact in planta with both Cf-4 and Ve1 and are required for the Cf-4– and Ve1-mediated hypersensitive response and immunity. Tomato SOBIR1/EVR interacts with most of the tested RLPs, but not with the RLKs FLS2, SERK1, SERK3a, BAK1, and CLV1. SOBIR1/EVR is required for stability of the Cf-4 and Ve1 receptors, supporting our observation that these RLPs are present in a complex with SOBIR1/EVR in planta. We show that SOBIR1/EVR is essential for RLP-mediated immunity and propose that the protein functions as a regulatory RLK of this type of cell-surface receptors.
How to effectively deploy plant resistances to pests and pathogens in crop breeding
Seifi Abdolabad, A.R. ; Visser, R.G.F. ; Bai, Y. - \ 2013
Euphytica 190 (2013)3. - ISSN 0014-2336 - p. 321 - 334.
durable disease resistance - systemic acquired-resistance - nbs-lrr genes - cell-death - arabidopsis-thaliana - confers resistance - fungal pathogen - small rnas - microbe interactions - nematode resistance
Food shortage is currently a serious worldwide problem, and the fact that it will become an even greater problem in the coming decades underscores the necessity for more translational research in plant sciences that is directed towards crop breeding. Biotic stresses are important determinants of reduced crop production and, therefore, a better understanding of the resistance mechanisms utilized by plants to survive and remain productive following attacks by pathogens and pests is a major research goal of plant scientists and crop breeders. During the last two decades tremendous progress has been made in this field. Here we first summarize recent findings on how plants respond to their pathogens and pests and then we discuss resistance mechanisms in three categories based on differences in spectrum, durability and evolution. We also review different breeding strategies that have been adopted on the basis of these findings for improving resistance to pests and pathogens in crops and evaluate these strategies for their sustainability
Tsw gene-based resistance is triggered by a functional RNA silencing suppressor protein of the Tomato spotted wilt virus
Ronde, D. de; Butterbach, P.B.E. ; Lohuis, H. ; Hedil, M. ; Lent, J.W.M. van; Kormelink, R.J.M. - \ 2013
Molecular Plant Pathology 14 (2013)4. - ISSN 1464-6722 - p. 405 - 415.
mediated plant transformation - capsicum-chinense - cell-death - disease-resistance - lycopersicon-esculentum - viral suppressors - sw-5 gene - potato - tospovirus - agrobacterium
As a result of contradictory reports, the avirulence (Avr) determinant that triggers Tsw gene-based resistance in Capsicum annuum against the Tomato spotted wilt virus (TSWV) is still unresolved. Here, the N and NSs genes of resistance-inducing (RI) and resistance-breaking (RB) isolates were cloned and transiently expressed in resistant Capsicum plants to determine the identity of the Avr protein. It was shown that the NSsRI protein triggered a hypersensitive response (HR) in Tsw-containing Capsicum plants, but not in susceptible Capsicum, whereas no HR was discerned after expression of the NRI/RB protein, or when NSsRB was expressed. Although NSsRI was able to suppress the silencing of a functional green fluorescence protein (GFP) construct during Agrobacterium tumefaciens transient assays on Nicotiana benthamiana, NSsRB had lost this capacity. The observation that RB isolates suppressed local GFP silencing during an infection indicated a recovery of RNA silencing suppressor activity for the NSs protein or the presence of another RNA interference (RNAi) suppressor. The role of NSs as RNA silencing suppressor and Avr determinant is discussed in the light of a putative interplay between RNAi and the natural Tsw resistance gene
Dual effects of N-acetyl-l-cysteine dependent on NQO1 activity: Suppressive or promotive of 9,10-phenanthrenequinone-induced toxicity
Toyooka, T. ; Shinmen, T. ; Aarts, J.M.M.J.G. ; Ibuki, Y. - \ 2012
Toxicology and Applied Pharmacology 264 (2012)3. - ISSN 0041-008X - p. 404 - 412.
double-strand breaks - yeast saccharomyces-cerevisiae - diesel exhaust particles - dt-diaphorase - nad(p)h-quinone oxidoreductase - oxidative stress - dna-damage - cell-death - quinone oxidoreductases - histone h2ax
A typical antioxidant, N-acetyl-L-cysteine (NAC) generally protects cells from oxidative damage induced by reactive oxygen species (ROS). 9,10-Phenanthrenequinone (9,10-PQ), a major quinone in diesel exhaust particles, produces ROS in redox cycling following two-electron reduction by NAD(P)H:quinone oxidoreductase 1 (NQO1), which has been considered as a cause of its cyto- and genotoxicity. In this study, we show that NAC unexpectedly augments the toxicity of 9,10-PQ in cells with low NQO1 activity. In four human skin cell lines, the expression and the activity of NQO1 were lower than in human adenocarcinoma cell lines, A549 and MCF7. In the skin cells, the cytotoxicity of 9,10-PQ was significantly enhanced by addition of NAC. The formation of DNA double strand breaks accompanying phosphorylation of histone H2AX, was also remarkably augmented. On the other hand, the cyto- and genotoxicity were suppressed by addition of NAC in the adenocarcinoma cells. Two contrasting experiments: overexpression of NQO1 in CHO-K1 cells which originally expressed low NQO1 levels, and knock-down of NQO1 in the adenocarcinoma cell line A549 by transfection of RNAi, also showed that NAC suppressed 9,10-PQ-induced toxicity in cell lines expressing high NQO1 activity and enhanced it in cell lines with low NQO1 activity. The results suggested that dual effects of NAC on the cyto- and genotoxicity of 9,10-PQ were dependent on tissue-specific NQO1 activity.
Plasma membrane calcium ATPases are important components of receptor-mediated signaling in plant immune responses and development.
Frei dit Frey, N. ; Mbengue, M. ; Kwaaitaal, M.A.C.J. ; Nitsch, L.M.C. ; Altenbach, D. ; Häweker, H. ; Lozano-Duran, R. ; Njo, M.F. ; Beeckman, T. ; Huettel, B. ; Borst, J.W. ; Panstruga, R. ; Robatzek, S. - \ 2012
Plant Physiology 159 (2012)2. - ISSN 0032-0889 - p. 798 - 809.
pattern-recognition receptors - arabidopsis-thaliana - innate immunity - molecular-patterns - protein-kinases - nadph oxidase - cell-death - defense - bak1 - brassinosteroids
Plasma membrane-resident receptor kinases (RKs) initiate signaling pathways important for plant immunity and development. In Arabidopsis (Arabidopsis thaliana), the receptor for the elicitor-active peptide epitope of bacterial flagellin, flg22, is encoded by FLAGELLIN SENSING2 (FLS2), which promotes plant immunity. Despite its relevance, the molecular components regulating FLS2-mediated signaling remain largely unknown. We show that plasma membrane ARABIDOPSIS-AUTOINHIBITED Ca2+-ATPase (ACA8) forms a complex with FLS2 in planta. ACA8 and its closest homolog ACA10 are required for limiting the growth of virulent bacteria. One of the earliest flg22 responses is the transient increase of cytosolic Ca2+ ions, which is crucial for many of the well-described downstream responses (e.g. generation of reactive oxygen species and the transcriptional activation of defense-associated genes). Mutant aca8 aca10 plants show decreased flg22-induced Ca2+ and reactive oxygen species bursts and exhibit altered transcriptional reprogramming. In particular, mitogen-activated protein kinase-dependent flg22-induced gene expression is elevated, whereas calcium-dependent protein kinase-dependent flg22-induced gene expression is reduced. These results demonstrate that the fine regulation of Ca2+ fluxes across the plasma membrane is critical for the coordination of the downstream microbe-associated molecular pattern responses and suggest a mechanistic link between the FLS2 receptor complex and signaling kinases via the secondary messenger Ca2+. ACA8 also interacts with other RKs such as BRI1 and CLV1 known to regulate plant development, and both aca8 and aca10 mutants show morphological phenotypes, suggesting additional roles for ACA8 and ACA10 in developmental processes. Thus, Ca2+ ATPases appear to represent general regulatory components of RK-mediated signaling pathways.
Host Protein BSL1 Associates with Phytophthora infestans RXLR Effector AVR2 and the Solanum demissum Immune Receptor R2 to Mediate Disease Resistance
Saunders, D.G.O. ; Breen, S. ; Win, J. ; Schornack, S. ; Hein, I. ; Bozkurt, T.O. ; Champouret, N. ; Vleeshouwers, V.G.A.A. ; Birch, P.R.J. ; Gilroy, E.M. ; Kamoun, S. - \ 2012
The Plant Cell 24 (2012)8. - ISSN 1040-4651 - p. 3420 - 3434.
nicotiana-benthamiana - plant transformation - avirulence genes - cell-death - arabidopsis - potato - activation - expression - virulence - target
Plant pathogens secrete effector proteins to modulate plant immunity and promote host colonization. Plant nucleotide binding leucine-rich repeat (NB-LRR) immunoreceptors recognize specific pathogen effectors directly or indirectly. Little is known about how NB-LRR proteins recognize effectors of filamentous plant pathogens, such as Phytophthora infestans. AVR2 belongs to a family of 13 sequence-divergent P. infestans RXLR effectors that are differentially recognized by members of the R2 NB-LRR family in Solanum demissum. We report that the putative plant phosphatase BSU-LIKE PROTEIN1 (BSL1) is required for R2-mediated perception of AVR2 and resistance to P. infestans. AVR2 associates with BSL1 and mediates the interaction of BSL1 with R2 in planta, possibly through the formation of a ternary complex. Strains of P. infestans that are virulent on R2 potatoes express an unrecognized form, Avr2-like (referred to as A2l). A2L can still interact with BSL1 but does not promote the association of BSL1 with R2. Our findings show that recognition of the P. infestans AVR2 effector by the NB-LRR protein R2 requires the putative phosphatase BSL1. This reveals that, similar to effectors of phytopathogenic bacteria, recognition of filamentous pathogen effectors can be mediated via a host protein that interacts with both the effector and the NB-LRR immunoreceptor.