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1.
New Phytol ; 2024 Jun 24.
Article in English | MEDLINE | ID: mdl-38922927

ABSTRACT

Leaf mould, caused by Fulvia fulva, is a devastating disease of tomato plants. In many commercial tomato cultivars, resistance to this disease is governed by the Cf-9 locus, which encodes five paralogous receptor-like proteins. Two of these proteins confer resistance: Cf-9C recognises the previously identified F. fulva effector Avr9 and provides resistance during all plant growth stages, while Cf-9B recognises the yet-unidentified F. fulva effector Avr9B and provides mature plant resistance only. In recent years, F. fulva strains have emerged that can overcome the Cf-9 locus, with Cf-9C circumvented through Avr9 deletion. To understand how Cf-9B is circumvented, we set out to identify Avr9B. Comparative genomics, transient expression assays and gene complementation experiments were used to identify Avr9B, while gene sequencing was used to assess Avr9B allelic variation across a world-wide strain collection. A strict correlation between Avr9 deletion and resistance-breaking mutations in Avr9B was observed in strains recently collected from Cf-9 cultivars, whereas Avr9 deletion but no mutations in Avr9B were observed in older strains. This research showcases how F. fulva has evolved to sequentially break down the Cf-9 locus and stresses the urgent need for commercial tomato cultivars that carry novel, stacked resistance genes active against this pathogen.

2.
Nat Commun ; 15(1): 4339, 2024 May 21.
Article in English | MEDLINE | ID: mdl-38773116

ABSTRACT

Cell-surface receptors form the front line of plant immunity. The leucine-rich repeat (LRR)-receptor-like kinases SOBIR1 and BAK1 are required for the functionality of the tomato LRR-receptor-like protein Cf-4, which detects the secreted effector Avr4 of the pathogenic fungus Fulvia fulva. Here, we show that the kinase domains of SOBIR1 and BAK1 directly phosphorylate each other and that residues Thr522 and Tyr469 of the kinase domain of Nicotiana benthamiana SOBIR1 are required for its kinase activity and for interacting with signalling partners, respectively. By knocking out multiple genes belonging to different receptor-like cytoplasmic kinase (RLCK)-VII subfamilies in N. benthamiana:Cf-4, we show that members of RLCK-VII-6, -7, and -8 differentially regulate the Avr4/Cf-4-triggered biphasic burst of reactive oxygen species. In addition, members of RLCK-VII-7 play an essential role in resistance against the oomycete pathogen Phytophthora palmivora. Our study provides molecular evidence for the specific roles of RLCKs downstream of SOBIR1/BAK1-containing immune complexes.


Subject(s)
Nicotiana , Plant Diseases , Plant Immunity , Plant Proteins , Protein Serine-Threonine Kinases , Nicotiana/immunology , Nicotiana/microbiology , Nicotiana/genetics , Nicotiana/metabolism , Plant Proteins/metabolism , Plant Proteins/genetics , Plant Immunity/genetics , Protein Serine-Threonine Kinases/metabolism , Protein Serine-Threonine Kinases/genetics , Plant Diseases/microbiology , Plant Diseases/immunology , Phytophthora/pathogenicity , Protein Kinases/metabolism , Protein Kinases/genetics , Phosphorylation , Gene Expression Regulation, Plant , Reactive Oxygen Species/metabolism , Signal Transduction
3.
Trends Plant Sci ; 2024 Apr 08.
Article in English | MEDLINE | ID: mdl-38594153

ABSTRACT

To resist biotic attacks, plants have evolved a sophisticated, receptor-based immune system. Cell-surface immune receptors, which are either receptor-like kinases (RLKs) or receptor-like proteins (RLPs), form the front line of the plant defense machinery. RLPs lack a cytoplasmic kinase domain for downstream immune signaling, and leucine-rich repeat (LRR)-containing RLPs constitutively associate with the RLK SOBIR1. The RLP/SOBIR1 complex was proposed to be the bimolecular equivalent of genuine RLKs. However, it appears that the molecular mechanisms by which RLP/SOBIR1 complexes and RLKs mount immunity show some striking differences. Here, we summarize the differences between RLP/SOBIR1 and RLK signaling, focusing on the way these receptors recruit the BAK1 co-receptor and elaborating on the negative crosstalk taking place between the two signaling networks.

4.
Plant Biotechnol J ; 22(7): 1913-1925, 2024 Jul.
Article in English | MEDLINE | ID: mdl-38366362

ABSTRACT

Potato is the third most important food crop worldwide. Potato production suffers from severe diseases caused by multiple detrimental plant pathogens, and broad-spectrum disease resistance genes are rarely identified in potato. Here we identified the potato non-specific lipid transfer protein StLTPa, which enhances species none-specific disease resistance against various pathogens, such as the oomycete pathogen Phytophthora infestans, the fungal pathogens Botrytis cinerea and Verticillium dahliae, and the bacterial pathogens Pectobacterium carotovorum and Ralstonia solanacearum. The StLTPa overexpression potato lines do not show growth penalty. Furthermore, we provide evidence that StLTPa binds to lipids present in the plasma membrane (PM) of the hyphal cells of P. infestans, leading to an increased permeability of the PM. Adding of PI(3,5)P2 and PI(3)P could compete the binding of StLTPa to pathogen PM and reduce the inhibition effect of StLTPa. The lipid-binding activity of StLTPa is essential for its role in pathogen inhibition and promotion of potato disease resistance. We propose that StLTPa enhances potato broad-spectrum disease resistance by binding to, and thereby promoting the permeability of the PM of the cells of various pathogens. Overall, our discovery illustrates that increasing the expression of a single gene in potato enhances potato disease resistance against different pathogens without growth penalty.


Subject(s)
Carrier Proteins , Cell Membrane , Disease Resistance , Phytophthora infestans , Plant Diseases , Plant Proteins , Solanum tuberosum , Solanum tuberosum/microbiology , Solanum tuberosum/genetics , Solanum tuberosum/metabolism , Solanum tuberosum/immunology , Disease Resistance/genetics , Plant Diseases/microbiology , Plant Diseases/immunology , Cell Membrane/metabolism , Plant Proteins/genetics , Plant Proteins/metabolism , Phytophthora infestans/pathogenicity , Carrier Proteins/metabolism , Carrier Proteins/genetics , Ralstonia solanacearum/pathogenicity , Ralstonia solanacearum/physiology , Botrytis , Plants, Genetically Modified , Pectobacterium carotovorum
5.
STAR Protoc ; 4(4): 102577, 2023 Dec 15.
Article in English | MEDLINE | ID: mdl-37733594

ABSTRACT

Protein-protein interactions (PPIs) in crop plants remain largely unexplored. Here, we provide a protocol for identifying PPIs in potato (Solanum tuberosum) using TurboID-mediated proximity labeling. We transiently expressed constructs for a nucleus-located transcription factor and a plasma membrane-localized receptor-like kinase fused to TurboID to identify PPIs in potato leaves. We describe the plasmid construction, plant material, agroinfiltration, biotin treatment, protein isolation, free biotin removal, western blot analysis, and enrichment of biotinylated proteins for mass spectrometry analysis.


Subject(s)
Protein Interaction Maps , Solanum tuberosum , Solanum tuberosum/genetics , Biotin , Plants , Transcription Factors
6.
Mol Plant Pathol ; 24(5): 399-412, 2023 05.
Article in English | MEDLINE | ID: mdl-36782107

ABSTRACT

Mitogen-activated protein kinase (MAPK) cascades play pivotal roles in plant responses to both biotic and abiotic stress. A screen of a Nicotiana benthamiana cDNA virus-induced gene silencing (VIGS) library for altered plant responses to inoculation with Phytophthora infestans previously identified an NbMKK gene, encoding a clade D MAPKK that we renamed as NbMKK5, which is involved in immunity to P. infestans. To study the role of the potato orthologous gene, referred to as StMKK5, in the response to P. infestans, we transiently overexpressed StMKK5 in N. benthamiana and observed that cell death occurred at 2 days postinfiltration. Silencing of the highly conserved eukaryotic protein SGT1 delayed the StMKK5-induced cell death, whereas silencing of the MAPK-encoding gene NbSIPK completely abolished the cell death response. Further investigations showed that StMKK5 interacts with, and directly phosphorylates, StSIPK. Furthermore, both StMKK5 and StSIPK trigger salicylic acid (SA)- and ethylene (Eth)-related gene expression, and co-expression of the salicylate hydroxylase NahG with the negative regulator of Eth signalling CTR1 hampers StSIPK-triggered cell death. This observation indicates that the cell death triggered by StMKK5-StSIPK is dependent on the combination of SA- and Eth-signalling. By introducing point mutations, we showed that the kinase activity of both StMKK5 and StSIPK is required for triggering cell death. Genetic analysis showed that StMKK5 depends on StSIPK to trigger plant resistance. Thus, our results define a potato StMKK5-SIPK module that positively regulates immunity to P. infestans via activation of both the SA and Eth signalling pathways.


Subject(s)
Phytophthora infestans , Solanum tuberosum , Salicylic Acid/metabolism , Plant Proteins/genetics , Plant Proteins/metabolism , Ethylenes/metabolism , Phytophthora infestans/physiology , Plant Diseases , Nicotiana/metabolism
7.
Mol Plant Pathol ; 24(5): 474-494, 2023 05.
Article in English | MEDLINE | ID: mdl-36790136

ABSTRACT

Fulvia fulva and Dothistroma septosporum are closely related apoplastic pathogens with similar lifestyles but different hosts: F. fulva is a pathogen of tomato, whilst D. septosporum is a pathogen of pine trees. In 2012, the first genome sequences of these pathogens were published, with F. fulva and D. septosporum having highly fragmented and near-complete assemblies, respectively. Since then, significant advances have been made in unravelling their genome architectures. For instance, the genome of F. fulva has now been assembled into 14 chromosomes, 13 of which have synteny with the 14 chromosomes of D. septosporum, suggesting these pathogens are even more closely related than originally thought. Considerable advances have also been made in the identification and functional characterization of virulence factors (e.g., effector proteins and secondary metabolites) from these pathogens, thereby providing new insights into how they promote host colonization or activate plant defence responses. For example, it has now been established that effector proteins from both F. fulva and D. septosporum interact with cell-surface immune receptors and co-receptors to activate the plant immune system. Progress has also been made in understanding how F. fulva and D. septosporum have evolved with their host plants, whilst intensive research into pandemics of Dothistroma needle blight in the Northern Hemisphere has shed light on the origins, migration, and genetic diversity of the global D. septosporum population. In this review, we specifically summarize advances made in our understanding of the F. fulva-tomato and D. septosporum-pine pathosystems over the last 10 years.


Subject(s)
Ascomycota , Cladosporium , Host Microbial Interactions , Pinus , Ascomycota/genetics , Cladosporium/genetics , Pinus/immunology , Pinus/microbiology , Genome, Fungal/genetics
8.
New Phytol ; 238(2): 781-797, 2023 04.
Article in English | MEDLINE | ID: mdl-36653957

ABSTRACT

Ubiquitin-like domain-containing proteins (UDPs) are involved in the ubiquitin-proteasome system because of their ability to interact with the 26S proteasome. Here, we identified potato StUDP as a target of the Phytophthora infestans RXLR effector Pi06432 (PITG_06432), which supresses the salicylic acid (SA)-related immune pathway. By overexpressing and silencing of StUDP in potato, we show that StUDP negatively regulates plant immunity against P. infestans. StUDP interacts with, and destabilizes, the 26S proteasome subunit that is referred to as REGULATORY PARTICLE TRIPLE-A ATP-ASE (RPT) subunit StRPT3b. This destabilization represses the proteasome activity. Proteomic analysis and Western blotting show that StUDP decreases the stability of the master transcription factor SYSTEMIC ACQUIRED RESISTANCE DEFICIENT 1 (SARD1) in SA biosynthesis. StUDP negatively regulates the SA signalling pathway by repressing the proteasome activity and destabilizing StSARD1, leading to a decreased expression of the SARD1-targeted gene ISOCHORISMATE SYNTHASE 1 and thereby a decrease in SA content. Pi06432 stabilizes StUDP, and it depends on StUDP to destabilize StRPT3b and thereby supress the proteasome activity. Our study reveals that the P. infestans effector Pi06432 targets StUDP to hamper the homeostasis of the proteasome by the degradation of the proteasome subunit StRPT3b and thereby suppresses SA-related immunity.


Subject(s)
Phytophthora infestans , Solanum tuberosum , Phytophthora infestans/metabolism , Ubiquitins/metabolism , Proteasome Endopeptidase Complex/metabolism , Proteomics , Plant Immunity , Plant Diseases , Plant Proteins/genetics , Plant Proteins/metabolism
9.
Plant Biotechnol J ; 21(3): 646-661, 2023 03.
Article in English | MEDLINE | ID: mdl-36519513

ABSTRACT

Phytophthora infestans causes severe losses in potato production. The MAPK kinase StMKK1 was previously found to negatively regulate potato immunity to P. infestans. Our results showed that StMKK1 interacts with a protein tyrosine phosphatase, referred to as StPTP1a, and StMKK1 directly phosphorylates StPTP1a at residues Ser-99, Tyr-223 and Thr-290. StPTP1a is a functional phosphatase and the phosphorylation of StPTP1a at these three residues enhances its stability and catalytic activity. StPTP1a negatively regulates potato immunity and represses SA-related gene expression. Furthermore, StPTP1a interacts with, and dephosphorylates, the StMKK1 downstream signalling targets StMPK4 and -7 at their Tyr-203 residue resulting in the repression of salicylic acid (SA)-related immunity. Silencing of NbPTP1a + NbMPK4 or NbPTP1a + NbMPK7 abolished the plant immunity to P. infestans caused by NbPTP1a silencing, indicating that PTP1a functions upstream of NbMPK4 and NbMPK7. StMKK1 requires StPTP1a to negatively regulate SA-related immunity and StPTP1a is phosphorylated and stabilized during immune activation to promote the de-phosphorylation of StMPK4 and -7. Our results reveal that potato StMKK1 activates and stabilizes the tyrosine phosphatase StPTP1a that in its turn de-phosphorylates StMPK4 and -7, thereby repressing plant SA-related immunity.


Subject(s)
Phytophthora infestans , Solanum tuberosum , Solanum tuberosum/genetics , Plant Proteins/genetics , Plant Immunity , Phytophthora infestans/physiology , Protein Tyrosine Phosphatases/metabolism , Plant Diseases/genetics
10.
New Phytol ; 236(6): 2249-2264, 2022 12.
Article in English | MEDLINE | ID: mdl-36151929

ABSTRACT

Heterodimeric complexes incorporating the lipase-like proteins EDS1 with PAD4 or SAG101 are central hubs in plant innate immunity. EDS1 functions encompass signal relay from TIR domain-containing intracellular NLR-type immune receptors (TNLs) towards RPW8-type helper NLRs (RNLs) and, in Arabidopsis thaliana, bolstering of signaling and resistance mediated by cell-surface pattern recognition receptors (PRRs). Increasing evidence points to the activation of EDS1 complexes by small molecule binding. We used CRISPR/Cas-generated mutant lines and agroinfiltration-based complementation assays to interrogate functions of EDS1 complexes in Nicotiana benthamiana. We did not detect impaired PRR signaling in N. benthamiana lines deficient in EDS1 complexes or RNLs. Intriguingly, in assays monitoring functions of SlEDS1-NbEDS1 complexes in N. benthamiana, mutations within the SlEDS1 catalytic triad could abolish or enhance TNL immunity. Furthermore, nuclear EDS1 accumulation was sufficient for N. benthamiana TNL (Roq1) immunity. Reinforcing PRR signaling in Arabidopsis might be a derived function of the TNL/EDS1 immune sector. Although Solanaceae EDS1 functionally depends on catalytic triad residues in some contexts, our data do not support binding of a TNL-derived small molecule in the triad environment. Whether and how nuclear EDS1 activity connects to membrane pore-forming RNLs remains unknown.


Subject(s)
Arabidopsis Proteins , Arabidopsis , Nicotiana/genetics , Nicotiana/metabolism , Arabidopsis Proteins/metabolism , DNA-Binding Proteins/metabolism , Plant Immunity/genetics , Arabidopsis/metabolism , Receptors, Cell Surface/metabolism , Plant Diseases , Carboxylic Ester Hydrolases/metabolism
11.
Mol Plant Pathol ; 23(3): 431-446, 2022 03.
Article in English | MEDLINE | ID: mdl-34913556

ABSTRACT

To identify host factors for tomato spotted wilt orthotospovirus (TSWV), a virus-induced gene silencing (VIGS) screen using tobacco rattle virus (TRV) was performed on Nicotiana benthamiana for TSWV susceptibility. To rule out any negative effect on the plants' performance due to a double viral infection, the method was optimized to allow screening of hundreds of clones in a standardized fashion. To normalize the results obtained in and between experiments, a set of controls was developed to evaluate in a consist manner both VIGS efficacy and the level of TSWV resistance. Using this method, 4532 random clones of an N. benthamiana cDNA library were tested, resulting in five TRV clones that provided nearly complete resistance against TSWV. Here we report on one of these clones, of which the insert targets a small gene family coding for the ribosomal protein S6 (RPS6) that is part of the 40S ribosomal subunit. This RPS6 family is represented by three gene clades in the genome of Solanaceae family members, which were jointly important for TSWV susceptibility. Interestingly, RPS6 is a known host factor implicated in the replication of different plant RNA viruses, including the negative-stranded TSWV and the positive-stranded potato virus X.


Subject(s)
RNA Viruses , Solanum lycopersicum , Tospovirus , Plant Diseases , Ribosomal Protein S6 , Nicotiana/genetics
12.
Viruses ; 13(11)2021 10 30.
Article in English | MEDLINE | ID: mdl-34834996

ABSTRACT

The tripartite genome of the negative-stranded RNA virus Tomato spotted wilt orthotospovirus (TSWV) is assembled, together with two viral proteins, the nucleocapsid protein and the RNA-dependent RNA polymerase, into infectious ribonucleoprotein complexes (RNPs). These two viral proteins are, together, essential for viral replication and transcription, yet our knowledge on the host factors supporting these two processes remains limited. To fill this knowledge gap, the protein composition of viral RNPs collected from TSWV-infected Nicotiana benthamiana plants, and of those collected from a reconstituted TSWV replicon system in the yeast Saccharomyces cerevisiae, was analysed. RNPs obtained from infected plant material were enriched for plant proteins implicated in (i) sugar and phosphate transport and (ii) responses to cellular stress. In contrast, the yeast-derived viral RNPs primarily contained proteins implicated in RNA processing and ribosome biogenesis. The latter suggests that, in yeast, the translational machinery is recruited to these viral RNPs. To examine whether one of these cellular proteins is important for a TSWV infection, the corresponding N. benthamiana genes were targeted for virus-induced gene silencing, and these plants were subsequently challenged with TSWV. This approach revealed four host factors that are important for systemic spread of TSWV and disease symptom development.


Subject(s)
Nicotiana/virology , Peptide Elongation Factor 1/metabolism , Protein Isoforms/metabolism , Tospovirus/physiology , Host Microbial Interactions/genetics , Host Microbial Interactions/physiology , Solanum lycopersicum , Nucleocapsid Proteins , Peptide Elongation Factor 1/genetics , Plant Diseases/virology , Plant Proteins/genetics , Plant Proteins/metabolism , Protein Isoforms/genetics , Replicon , Ribonucleoproteins/metabolism , Tospovirus/genetics , Viral Proteins/genetics , Viral Proteins/metabolism , Virus Replication
13.
Nature ; 598(7881): 495-499, 2021 10.
Article in English | MEDLINE | ID: mdl-34497423

ABSTRACT

Plants deploy cell-surface and intracellular leucine rich-repeat domain (LRR) immune receptors to detect pathogens1. LRR receptor kinases and LRR receptor proteins at the plasma membrane recognize microorganism-derived molecules to elicit pattern-triggered immunity (PTI), whereas nucleotide-binding LRR proteins detect microbial effectors inside cells to confer effector-triggered immunity (ETI). Although PTI and ETI are initiated in different host cell compartments, they rely on the transcriptional activation of similar sets of genes2, suggesting pathway convergence upstream of nuclear events. Here we report that PTI triggered by the Arabidopsis LRR receptor protein RLP23 requires signalling-competent dimers of the lipase-like proteins EDS1 and PAD4, and of ADR1 family helper nucleotide-binding LRRs, which are all components of ETI. The cell-surface LRR receptor kinase SOBIR1 links RLP23 with EDS1, PAD4 and ADR1 proteins, suggesting the formation of supramolecular complexes containing PTI receptors and transducers at the inner side of the plasma membrane. We detected similar evolutionary patterns in LRR receptor protein and nucleotide-binding LRR genes across Arabidopsis accessions; overall higher levels of variation in LRR receptor proteins than in LRR receptor kinases are consistent with distinct roles of these two receptor families in plant immunity. We propose that the EDS1-PAD4-ADR1 node is a convergence point for defence signalling cascades, activated by both surface-resident and intracellular LRR receptors, in conferring pathogen immunity.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/immunology , Carboxylic Ester Hydrolases/metabolism , DNA-Binding Proteins/metabolism , Plant Immunity , Protein Serine-Threonine Kinases/metabolism , Arabidopsis Proteins/chemistry , Carboxylic Ester Hydrolases/chemistry , DNA-Binding Proteins/chemistry , Protein Domains , Protein Kinases/chemistry , Protein Kinases/metabolism , Protein Multimerization , Protein Serine-Threonine Kinases/chemistry , Receptors, Cell Surface/chemistry , Receptors, Cell Surface/metabolism
14.
Mol Plant Pathol ; 22(6): 644-657, 2021 06.
Article in English | MEDLINE | ID: mdl-33764635

ABSTRACT

A cascade formed by phosphorylation events of mitogen-activated protein kinases (MAPKs) takes part in plant stress responses. However, the roles of these MAPKs in resistance of potato (Solanum tuberosum) against Phytophthora pathogens is not well studied. Our previous work showed that a Phytophthora infestans RXLR effector targets and stabilizes the negative regulator of MAPK kinase 1 of potato (StMKK1). Because in Arabidopsis thaliana the AtMPK4 is the downstream phosphorylation target of AtMKK1, we performed a phylogenetic analysis and found that potato StMPK4/6/7 are closely related and are orthologs of AtMPK4/5/11/12. Overexpression of StMPK4/7 enhances plant resistance to P. infestans and P. parasitica. Yeast two-hybrid analysis revealed that StMPK7 interacts with StMKK1, and StMPK7 is phosphorylated on flg22 treatment and by expressing constitutively active StMKK1 (CA-StMKK1), indicating that StMPK7 is a direct downstream signalling partner of StMKK1. Overexpression of StMPK7 in potato enhances potato resistance to P. infestans. Constitutively active StMPK7 (CA-StMPK7; StMPK7D198G, E202A ) was found to promote immunity to Phytophthora pathogens and to trigger host cell death when overexpressed in Nicotiana benthamiana leaves. Cell death triggered by CA-StMPK7 is SGT1/RAR1-dependent. Furthermore, cell death triggered by CA-StMPK7 is suppressed on coexpression with the salicylate hydroxylase NahG, and StMPK7 activation promotes salicylic acid (SA)-responsive gene expression. We conclude that potato StMPK7 is a downstream signalling component of the phosphorelay cascade involving StMKK1 and StMPK7 plays a role in immunity to Phytophthora pathogens via an SA-dependent signalling pathway.


Subject(s)
Disease Resistance , MAP Kinase Signaling System , Mitogen-Activated Protein Kinases/metabolism , Phytophthora infestans/physiology , Plant Diseases/immunology , Solanum tuberosum/genetics , Cell Death , Gene Expression , Mitogen-Activated Protein Kinases/genetics , Phylogeny , Plant Diseases/parasitology , Plant Leaves/genetics , Plant Leaves/immunology , Plant Leaves/parasitology , Plant Proteins/genetics , Plant Proteins/metabolism , Salicylic Acid/metabolism , Solanum tuberosum/immunology , Solanum tuberosum/parasitology , Nicotiana/genetics , Nicotiana/immunology , Nicotiana/parasitology
16.
Mol Plant Pathol ; 22(3): 361-372, 2021 03.
Article in English | MEDLINE | ID: mdl-33497519

ABSTRACT

Studies on plant-pathogen interactions often involve monitoring disease symptoms or responses of the host plant to pathogen-derived immunogenic patterns, either visually or by staining the plant tissue. Both these methods have limitations with respect to resolution, reproducibility, and the ability to quantify the results. In this study we show that red light detection by the red fluorescent protein (RFP) channel of a multipurpose fluorescence imaging system that is probably available in many laboratories can be used to visualize plant tissue undergoing cell death. Red light emission is the result of chlorophyll fluorescence on thylakoid membrane disassembly during the development of a programmed cell death process. The activation of programmed cell death can occur during either a hypersensitive response to a biotrophic pathogen or an apoptotic cell death triggered by a necrotrophic pathogen. Quantifying the intensity of the red light signal enables the magnitude of programmed cell death to be evaluated and provides a readout of the plant immune response in a faster, safer, and nondestructive manner when compared to previously developed chemical staining methodologies. This application can be implemented to screen for differences in symptom severity in plant-pathogen interactions, and to visualize and quantify in a more sensitive and objective manner the intensity of the plant response on perception of a given immunological pattern. We illustrate the utility and versatility of the method using diverse immunogenic patterns and pathogens.


Subject(s)
Apoptosis , Arabidopsis/physiology , Host-Pathogen Interactions , Lilium/physiology , Nicotiana/physiology , Arabidopsis/cytology , Arabidopsis/immunology , Arabidopsis/microbiology , Light , Lilium/genetics , Lilium/immunology , Lilium/microbiology , Optical Imaging , Plant Leaves/immunology , Plant Leaves/microbiology , Plant Leaves/radiation effects , Reproducibility of Results , Nicotiana/immunology , Nicotiana/microbiology , Nicotiana/radiation effects
17.
Proc Biol Sci ; 287(1941): 20202723, 2020 12 23.
Article in English | MEDLINE | ID: mdl-33352079

ABSTRACT

Natural plant populations encounter strong pathogen pressure and defence-associated genes are known to be under selection dependent on the pressure by the pathogens. Here, we use populations of the wild tomato Solanum chilense to investigate natural resistance against Cladosporium fulvum, a well-known ascomycete pathogen of domesticated tomatoes. Host populations used are from distinct geographical origins and share a defined evolutionary history. We show that distinct populations of S. chilense differ in resistance against the pathogen. Screening for major resistance gene-mediated pathogen recognition throughout the whole species showed clear geographical differences between populations and complete loss of pathogen recognition in the south of the species range. In addition, we observed high complexity in a homologues of Cladosporium resistance (Hcr) locus, underlying the recognition of C. fulvum, in central and northern populations. Our findings show that major gene-mediated recognition specificity is diverse in a natural plant-pathosystem. We place major gene resistance in a geographical context that also defined the evolutionary history of that species. Data suggest that the underlying loci are more complex than previously anticipated, with small-scale gene recombination being possibly responsible for maintaining balanced polymorphisms in the populations that experience pathogen pressure.


Subject(s)
Ascomycota , Plant Diseases/microbiology , Solanum lycopersicum/physiology , Cladosporium , Disease Resistance , Genes, Plant , Solanum lycopersicum/microbiology , Solanum
18.
Trends Plant Sci ; 24(7): 587-601, 2019 07.
Article in English | MEDLINE | ID: mdl-31171472

ABSTRACT

Models are extensively used to describe the coevolution of plants and microbial attackers. Such models distinguish between different classes of plant immune responses, based on the type of danger signal that is recognized or on the strength of the defense response that the danger signal provokes. However, recent molecular and biochemical advances have shown that these dichotomies are blurred. With molecular proof in hand, we propose here to abandon the current classification of plant immune responses, and to define the different forms of plant immunity solely based on the site of microbe recognition - either extracellular or intracellular. Using this spatial partition, our 'spatial immunity model' facilitates a broadly inclusive, but clearly distinguishing nomenclature to describe immune signaling in plant-microbe interactions.


Subject(s)
Plant Diseases , Plant Immunity , Plants , Signal Transduction
19.
Mol Plant Pathol ; 20(6): 751-764, 2019 06.
Article in English | MEDLINE | ID: mdl-30938041

ABSTRACT

The transfer of well-studied native and chimeric pattern recognition receptors (PRRs) to susceptible plants is a proven strategy to improve host resistance. In most cases, the ectodomain determines PRR recognition specificity, while the endodomain determines the intensity of the immune response. Here we report the generation and characterization of the chimeric receptor EFR-Cf-9, which carries the ectodomain of the Arabidopsis thaliana EF-Tu receptor (EFR) and the endodomain of the tomato Cf-9 resistance protein. Both transient and stable expression of EFR-Cf-9 triggered a robust hypersensitive response (HR) upon elf18 treatment in tobacco. Co-immunoprecipitation and virus-induced gene silencing studies showed that EFR-Cf-9 constitutively interacts with SUPPRESSOR OF BIR1-1 (SOBIR1) co-receptor, and requires both SOBIR1 and kinase-active BRI1-ASSOCIATED KINASE1 (BAK1) for its function. Transgenic plants expressing EFR-Cf-9 were more resistant to the (hemi)biotrophic bacterial pathogens Pseudomonas amygdali pv. tabaci (Pta) 11528 and Pseudomonas syringae pv. tomato DC3000, and mounted an HR in response to high doses of Pta 11528 and P. carotovorum. Taken together, these data indicate that the EFR-Cf-9 chimera is a valuable tool for both investigating the molecular mechanisms responsible for the activation of defence responses by PRRs, and for potential biotechnological use to improve crop disease resistance.


Subject(s)
Plant Proteins/metabolism , Plants, Genetically Modified/immunology , Plants, Genetically Modified/microbiology , Arabidopsis/immunology , Arabidopsis/metabolism , Arabidopsis/microbiology , Disease Resistance/genetics , Disease Resistance/physiology , Gene Expression Regulation, Plant , Solanum lycopersicum/immunology , Solanum lycopersicum/metabolism , Solanum lycopersicum/microbiology , Peptide Elongation Factor Tu/genetics , Peptide Elongation Factor Tu/metabolism , Plant Diseases/microbiology , Plant Immunity/genetics , Plant Immunity/physiology , Plant Proteins/genetics , Plants, Genetically Modified/metabolism , Receptors, Pattern Recognition/immunology , Receptors, Pattern Recognition/metabolism , Signal Transduction/genetics , Signal Transduction/physiology , Nicotiana/immunology , Nicotiana/metabolism , Nicotiana/microbiology
20.
Mol Plant Pathol ; 20(3): 410-422, 2019 03.
Article in English | MEDLINE | ID: mdl-30407725

ABSTRACT

Leucine-rich repeat-receptor-like proteins (LRR-RLPs) and LRR-receptor-like kinases (LRR-RLKs) trigger immune signalling to promote plant resistance against pathogens. LRR-RLPs lack an intracellular kinase domain, and several of these receptors have been shown to constitutively interact with the LRR-RLK Suppressor of BIR1-1/EVERSHED (SOBIR1/EVR) to form signalling-competent receptor complexes. Ligand perception by LRR-RLPs initiates recruitment of the co-receptor BRI1-Associated Kinase 1/Somatic Embryogenesis Receptor Kinase 3 (BAK1/SERK3) to the LRR-RLP/SOBIR1 complex, thereby activating LRR-RLP-mediated immunity. We employed phosphorylation analysis of in planta-produced proteins, live cell imaging, gene silencing and co-immunoprecipitation to investigate the roles of SOBIR1 and BAK1 in immune signalling. We show that Arabidopsis thaliana (At) SOBIR1, which constitutively activates immune responses when overexpressed in planta, is highly phosphorylated. Moreover, in addition to the kinase activity of SOBIR1 itself, kinase-active BAK1 is essential for AtSOBIR1-induced constitutive immunity and for the phosphorylation of AtSOBIR1. Furthermore, the defence response triggered by the tomato LRR-RLP Cf-4 on perception of Avr4 from the extracellular pathogenic fungus Cladosporium fulvum is dependent on kinase-active BAK1. We argue that, in addition to the trans-autophosphorylation of SOBIR1, it is likely that SOBIR1 and BAK1 transphosphorylate, and thereby activate the receptor complex. The signalling-competent cell surface receptor complex subsequently activates downstream cytoplasmic signalling partners to initiate RLP-mediated immunity.


Subject(s)
Arabidopsis Proteins/metabolism , Plant Immunity/physiology , Protein Kinases/metabolism , Protein Serine-Threonine Kinases/metabolism , Gene Expression Regulation, Plant , Phosphorylation/genetics , Phosphorylation/physiology , Plant Immunity/genetics , Plants, Genetically Modified , Signal Transduction/genetics , Signal Transduction/physiology
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