ABSTRACT
Nucleotide-binding, leucine-rich repeat receptors (NLRs) are major immune receptors in plants and animals. Upon activation, the Arabidopsis NLR protein ZAR1 forms a pentameric resistosome in vitro and triggers immune responses and cell death in plants. In this study, we employed single-molecule imaging to show that the activated ZAR1 protein can form pentameric complexes in the plasma membrane. The ZAR1 resistosome displayed ion channel activity in Xenopus oocytes in a manner dependent on a conserved acidic residue Glu11 situated in the channel pore. Pre-assembled ZAR1 resistosome was readily incorporated into planar lipid-bilayers and displayed calcium-permeable cation-selective channel activity. Furthermore, we show that activation of ZAR1 in the plant cell led to Glu11-dependent Ca2+ influx, perturbation of subcellular structures, production of reactive oxygen species, and cell death. The results thus support that the ZAR1 resistosome acts as a calcium-permeable cation channel to trigger immunity and cell death.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/immunology , Arabidopsis/metabolism , Calcium/metabolism , Carrier Proteins/metabolism , Disease Resistance/immunology , Plant Immunity , Signal Transduction , Animals , Cell Death , Cell Membrane/metabolism , Cell Membrane Permeability , Glutamic Acid/metabolism , Lipid Bilayers/metabolism , Oocytes/metabolism , Plant Cells/metabolism , Protein Multimerization , Protoplasts/metabolism , Reactive Oxygen Species/metabolism , Single Molecule Imaging , Vacuoles/metabolism , XenopusABSTRACT
In plants, pathogen effector-triggered immunity (ETI) often leads to programmed cell death, which is restricted by NPR1, an activator of systemic acquired resistance. However, the biochemical activities of NPR1 enabling it to promote defense and restrict cell death remain unclear. Here we show that NPR1 promotes cell survival by targeting substrates for ubiquitination and degradation through formation of salicylic acid-induced NPR1 condensates (SINCs). SINCs are enriched with stress response proteins, including nucleotide-binding leucine-rich repeat immune receptors, oxidative and DNA damage response proteins, and protein quality control machineries. Transition of NPR1 into condensates is required for formation of the NPR1-Cullin 3 E3 ligase complex to ubiquitinate SINC-localized substrates, such as EDS1 and specific WRKY transcription factors, and promote cell survival during ETI. Our analysis of SINCs suggests that NPR1 is centrally integrated into the cell death or survival decisions in plant immunity by modulating multiple stress-responsive processes in this quasi-organelle.
Subject(s)
Arabidopsis Proteins/immunology , Arabidopsis Proteins/metabolism , Cell Survival/immunology , Plant Immunity/immunology , Arabidopsis/immunology , Arabidopsis/metabolism , Gene Expression Regulation, Plant/immunology , Salicylic Acid/immunology , Salicylic Acid/metabolism , Ubiquitination/immunologyABSTRACT
Recognition of microbe-associated molecular patterns (MAMPs) is crucial for the plant's immune response. How this sophisticated perception system can be usefully deployed in roots, continuously exposed to microbes, remains a mystery. By analyzing MAMP receptor expression and response at cellular resolution in Arabidopsis, we observed that differentiated outer cell layers show low expression of pattern-recognition receptors (PRRs) and lack MAMP responsiveness. Yet, these cells can be gated to become responsive by neighbor cell damage. Laser ablation of small cell clusters strongly upregulates PRR expression in their vicinity, and elevated receptor expression is sufficient to induce responsiveness in non-responsive cells. Finally, localized damage also leads to immune responses to otherwise non-immunogenic, beneficial bacteria. Damage-gating is overridden by receptor overexpression, which antagonizes colonization. Our findings that cellular damage can "switch on" local immune responses helps to conceptualize how MAMP perception can be used despite the presence of microbial patterns in the soil.
Subject(s)
Arabidopsis/immunology , Plant Diseases/immunology , Plant Diseases/microbiology , Plant Roots/immunology , Receptors, Pattern Recognition/metabolism , Arabidopsis/growth & development , Arabidopsis/microbiology , Arabidopsis/radiation effects , Arabidopsis Proteins/metabolism , Arabidopsis Proteins/radiation effects , Ascorbate Peroxidases/metabolism , Ascorbate Peroxidases/radiation effects , Flagellin/pharmacology , Gene Expression Regulation, Plant/drug effects , Gene Expression Regulation, Plant/radiation effects , Laser Therapy/methods , Membrane Proteins/metabolism , Membrane Proteins/radiation effects , Microscopy, Confocal , Plant Roots/growth & development , Plant Roots/microbiology , Plant Roots/radiation effects , Protein Kinases/metabolism , Protein Kinases/radiation effects , Receptors, Pattern Recognition/radiation effects , Signal Transduction/drug effects , Signal Transduction/radiation effects , Time-Lapse ImagingABSTRACT
The molecular chaperone HSP90 facilitates the folding of several client proteins, including innate immune receptors and protein kinases. HSP90 is an essential component of plant and animal immunity, yet pathogenic strategies that directly target the chaperone have not been described. Here, we identify the HopBF1 family of bacterial effectors as eukaryotic-specific HSP90 protein kinases. HopBF1 adopts a minimal protein kinase fold that is recognized by HSP90 as a host client. As a result, HopBF1 phosphorylates HSP90 to completely inhibit the chaperone's ATPase activity. We demonstrate that phosphorylation of HSP90 prevents activation of immune receptors that trigger the hypersensitive response in plants. Consequently, HopBF1-dependent phosphorylation of HSP90 is sufficient to induce severe disease symptoms in plants infected with the bacterial pathogen, Pseudomonas syringae. Collectively, our results uncover a family of bacterial effector kinases with toxin-like properties and reveal a previously unrecognized betrayal mechanism by which bacterial pathogens modulate host immunity.
Subject(s)
Arabidopsis Proteins/metabolism , Bacterial Proteins/metabolism , HSP90 Heat-Shock Proteins/metabolism , Molecular Mimicry/immunology , Plant Immunity/physiology , Adenosine Triphosphatases/metabolism , Arabidopsis/immunology , Arabidopsis/metabolism , Arabidopsis/microbiology , Bacterial Proteins/chemistry , HEK293 Cells , HSP90 Heat-Shock Proteins/chemistry , HeLa Cells , Host Microbial Interactions/immunology , Humans , Phosphorylation , Plasmids/genetics , Protein Binding , Protein Folding , Protein Kinases/metabolism , Pseudomonas syringae/metabolism , Saccharomyces cerevisiae/metabolismABSTRACT
Following a previous microbial inoculation, plants can induce broad-spectrum immunity to pathogen infection, a phenomenon known as systemic acquired resistance (SAR). SAR establishment in Arabidopsis thaliana is regulated by the Lys catabolite pipecolic acid (Pip) and flavin-dependent-monooxygenase1 (FMO1). Here, we show that elevated Pip is sufficient to induce an FMO1-dependent transcriptional reprogramming of leaves that is reminiscent of SAR. In planta and in vitro analyses demonstrate that FMO1 functions as a pipecolate N-hydroxylase, catalyzing the biochemical conversion of Pip to N-hydroxypipecolic acid (NHP). NHP systemically accumulates in plants after microbial attack. When exogenously applied, it overrides the defect of NHP-deficient fmo1 in acquired resistance and acts as a potent inducer of plant immunity to bacterial and oomycete infection. Our work has identified a pathogen-inducible L-Lys catabolic pathway in plants that generates the N-hydroxylated amino acid NHP as a critical regulator of systemic acquired resistance to pathogen infection.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , Oxygenases/metabolism , Pipecolic Acids/metabolism , Plant Immunity/drug effects , Arabidopsis/enzymology , Arabidopsis/immunology , Arabidopsis Proteins/genetics , Gas Chromatography-Mass Spectrometry , Lysine/metabolism , Oomycetes/pathogenicity , Oxygenases/genetics , Pipecolic Acids/analysis , Pipecolic Acids/pharmacology , Plant Leaves/enzymology , Plant Leaves/immunology , Plant Leaves/metabolism , Pseudomonas syringae/pathogenicity , Transaminases/genetics , Transaminases/metabolismABSTRACT
Nuclear transport of immune receptors, signal transducers, and transcription factors is an essential regulatory mechanism for immune activation. Whether and how this process is regulated at the level of the nuclear pore complex (NPC) remains unclear. Here, we report that CPR5, which plays a key inhibitory role in effector-triggered immunity (ETI) and programmed cell death (PCD) in plants, is a novel transmembrane nucleoporin. CPR5 associates with anchors of the NPC selective barrier to constrain nuclear access of signaling cargos and sequesters cyclin-dependent kinase inhibitors (CKIs) involved in ETI signal transduction. Upon activation by immunoreceptors, CPR5 undergoes an oligomer to monomer conformational switch, which coordinates CKI release for ETI signaling and reconfigures the selective barrier to allow significant influx of nuclear signaling cargos through the NPC. Consequently, these coordinated NPC actions result in simultaneous activation of diverse stress-related signaling pathways and constitute an essential regulatory mechanism specific for ETI/PCD induction.
Subject(s)
Active Transport, Cell Nucleus/immunology , Arabidopsis Proteins/metabolism , Arabidopsis/immunology , Membrane Proteins/metabolism , Nuclear Pore/immunology , Signal Transduction , Arabidopsis/genetics , Arabidopsis Proteins/chemistry , Arabidopsis Proteins/genetics , Cell Cycle Proteins/metabolism , Gene Expression Regulation, Plant , Membrane Proteins/chemistry , Membrane Proteins/genetics , Protein ConformationABSTRACT
Microbial pathogens infect host cells by delivering virulence factors (effectors) that interfere with defenses. In plants, intracellular nucleotide-binding/leucine-rich repeat receptors (NLRs) detect specific effector interference and trigger immunity by an unknown mechanism. The Arabidopsis-interacting NLR pair, RRS1-R with RPS4, confers resistance to different pathogens, including Ralstonia solanacearum bacteria expressing the acetyltransferase effector PopP2. We show that PopP2 directly acetylates a key lysine within an additional C-terminal WRKY transcription factor domain of RRS1-R that binds DNA. This disrupts RRS1-R DNA association and activates RPS4-dependent immunity. PopP2 uses the same lysine acetylation strategy to target multiple defense-promoting WRKY transcription factors, causing loss of WRKY-DNA binding and transactivating functions needed for defense gene expression and disease resistance. Thus, RRS1-R integrates an effector target with an NLR complex at the DNA to switch a potent bacterial virulence activity into defense gene activation.
Subject(s)
Arabidopsis/immunology , Acetyltransferases/metabolism , Arabidopsis/microbiology , Arabidopsis Proteins/metabolism , DNA/metabolism , Models, Molecular , Plant Proteins/metabolism , Ralstonia solanacearum/enzymology , Ralstonia solanacearum/metabolism , Ralstonia solanacearum/pathogenicity , Transcription Factors/metabolismABSTRACT
Defense against pathogens in multicellular eukaryotes depends on intracellular immune receptors, yet surveillance by these receptors is poorly understood. Several plant nucleotide-binding, leucine-rich repeat (NB-LRR) immune receptors carry fusions with other protein domains. The Arabidopsis RRS1-R NB-LRR protein carries a C-terminal WRKY DNA binding domain and forms a receptor complex with RPS4, another NB-LRR protein. This complex detects the bacterial effectors AvrRps4 or PopP2 and then activates defense. Both bacterial proteins interact with the RRS1 WRKY domain, and PopP2 acetylates lysines to block DNA binding. PopP2 and AvrRps4 interact with other WRKY domain-containing proteins, suggesting these effectors interfere with WRKY transcription factor-dependent defense, and RPS4/RRS1 has integrated a "decoy" domain that enables detection of effectors that target WRKY proteins. We propose that NB-LRR receptor pairs, one member of which carries an additional protein domain, enable perception of pathogen effectors whose function is to target that domain.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/immunology , Plant Proteins/metabolism , Amino Acid Sequence , Arabidopsis/microbiology , Arabidopsis Proteins/chemistry , Bacterial Proteins/immunology , Immunity, Innate , Molecular Sequence Data , Protein Structure, Tertiary , Pseudomonas fluorescens/metabolism , Pseudomonas fluorescens/pathogenicity , Pseudomonas syringae/immunology , Pseudomonas syringae/metabolism , Nicotiana/immunology , Nicotiana/microbiologyABSTRACT
A mechanistic understanding of how plant pathogens modulate their hosts is critical for rationally engineered disease resistance in agricultural systems. Two new studies show that genomically paired plant immune receptors have incorporated decoy domains that structurally mimic pathogen virulence targets to monitor attempted host immunosuppression.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/immunology , Plant Proteins/metabolismABSTRACT
Calcium (Ca2+) is an essential nutrient for plants and a cellular signal, but excessive levels can be toxic and inhibit growth1,2. To thrive in dynamic environments, plants must monitor and maintain cytosolic Ca2+ homeostasis by regulating numerous Ca2+ transporters3. Here we report two signalling pathways in Arabidopsis thaliana that converge on the activation of vacuolar Ca2+/H+ exchangers (CAXs) to scavenge excess cytosolic Ca2+ in plants. One mechanism, activated in response to an elevated external Ca2+ level, entails calcineurin B-like (CBL) Ca2+ sensors and CBL-interacting protein kinases (CIPKs), which activate CAXs by phosphorylating a serine (S) cluster in the auto-inhibitory domain. The second pathway, triggered by molecular patterns associated with microorganisms, engages the immune receptor complex FLS2-BAK1 and the associated cytoplasmic kinases BIK1 and PBL1, which phosphorylate the same S-cluster in CAXs to modulate Ca2+ signals in immunity. These Ca2+-dependent (CBL-CIPK) and Ca2+-independent (FLS2-BAK1-BIK1/PBL1) mechanisms combine to balance plant growth and immunity by regulating cytosolic Ca2+ homeostasis.
Subject(s)
Arabidopsis , Calcium , Homeostasis , Plant Immunity , Arabidopsis/cytology , Arabidopsis/growth & development , Arabidopsis/immunology , Arabidopsis/metabolism , Arabidopsis Proteins/metabolism , Calcium/metabolism , Calcium-Binding Proteins/metabolism , Cytosol/metabolism , Phosphorylation , Phosphoserine/metabolism , Protein Serine-Threonine Kinases/metabolism , Cation Transport Proteins/metabolism , Antiporters/metabolismABSTRACT
Iron is critical during host-microorganism interactions1-4. Restriction of available iron by the host during infection is an important defence strategy, described as nutritional immunity5. However, this poses a conundrum for externally facing, absorptive tissues such as the gut epithelium or the plant root epidermis that generate environments that favour iron bioavailability. For example, plant roots acquire iron mostly from the soil and, when iron deficient, increase iron availability through mechanisms that include rhizosphere acidification and secretion of iron chelators6-9. Yet, the elevated iron bioavailability would also be beneficial for the growth of bacteria that threaten plant health. Here we report that microorganism-associated molecular patterns such as flagellin lead to suppression of root iron acquisition through a localized degradation of the systemic iron-deficiency signalling peptide Iron Man 1 (IMA1) in Arabidopsis thaliana. This response is also elicited when bacteria enter root tissues, but not when they dwell on the outer root surface. IMA1 itself has a role in modulating immunity in root and shoot, affecting the levels of root colonization and the resistance to a bacterial foliar pathogen. Our findings reveal an adaptive molecular mechanism of nutritional immunity that affects iron bioavailability and uptake, as well as immune responses.
Subject(s)
Arabidopsis Proteins , Arabidopsis , Bacteria , Intracellular Signaling Peptides and Proteins , Iron , Pathogen-Associated Molecular Pattern Molecules , Plant Roots , Arabidopsis/immunology , Arabidopsis/metabolism , Arabidopsis/microbiology , Arabidopsis Proteins/metabolism , Bacteria/immunology , Bacteria/metabolism , Flagellin/immunology , Gene Expression Regulation, Plant , Intracellular Signaling Peptides and Proteins/metabolism , Iron/metabolism , Plant Immunity , Plant Roots/immunology , Plant Roots/metabolism , Plant Roots/microbiology , Plant Shoots/immunology , Plant Shoots/metabolism , Plant Shoots/microbiology , Rhizosphere , Pathogen-Associated Molecular Pattern Molecules/immunology , Pathogen-Associated Molecular Pattern Molecules/metabolismABSTRACT
Plant nucleotide-binding leucine-rich repeat (NLR) immune receptors with an N-terminal Toll/interleukin-1 receptor (TIR) domain mediate recognition of strain-specific pathogen effectors, typically via their C-terminal ligand-sensing domains1. Effector binding enables TIR-encoded enzymatic activities that are required for TIR-NLR (TNL)-mediated immunity2,3. Many truncated TNL proteins lack effector-sensing domains but retain similar enzymatic and immune activities4,5. The mechanism underlying the activation of these TIR domain proteins remain unclear. Here we show that binding of the TIR substrates NAD+ and ATP induces phase separation of TIR domain proteins in vitro. A similar condensation occurs with a TIR domain protein expressed via its native promoter in response to pathogen inoculation in planta. The formation of TIR condensates is mediated by conserved self-association interfaces and a predicted intrinsically disordered loop region of TIRs. Mutations that disrupt TIR condensates impair the cell death activity of TIR domain proteins. Our data reveal phase separation as a mechanism for the activation of TIR domain proteins and provide insight into substrate-induced autonomous activation of TIR signalling to confer plant immunity.
Subject(s)
Adenosine Triphosphate , Arabidopsis , NAD , Nicotiana , Phase Separation , Plant Proteins , Protein Domains , Adenosine Triphosphate/metabolism , Arabidopsis/genetics , Arabidopsis/immunology , Arabidopsis/metabolism , Arabidopsis Proteins/chemistry , Arabidopsis Proteins/genetics , Arabidopsis Proteins/immunology , Arabidopsis Proteins/metabolism , Cell Death , Mutation , NAD/metabolism , Nicotiana/genetics , Nicotiana/immunology , Nicotiana/metabolism , NLR Proteins/chemistry , NLR Proteins/genetics , NLR Proteins/immunology , NLR Proteins/metabolism , Plant Diseases/immunology , Plant Immunity/genetics , Plant Proteins/chemistry , Plant Proteins/genetics , Plant Proteins/immunology , Plant Proteins/metabolism , Promoter Regions, Genetic , Protein Domains/genetics , Receptors, Immunologic/chemistry , Receptors, Immunologic/genetics , Receptors, Immunologic/immunology , Receptors, Immunologic/metabolism , Signal Transduction , Toll-Like Receptors/chemistry , Receptors, Interleukin-1/chemistryABSTRACT
Intraspecific genetic incompatibilities prevent the assembly of specific alleles into single genotypes and influence genome- and species-wide patterns of sequence variation. A common incompatibility in plants is hybrid necrosis, characterized by autoimmune responses due to epistatic interactions between natural genetic variants. By systematically testing thousands of F1 hybrids of Arabidopsis thaliana strains, we identified a small number of incompatibility hot spots in the genome, often in regions densely populated by nucleotide-binding domain and leucine-rich repeat (NLR) immune receptor genes. In several cases, these immune receptor loci interact with each other, suggestive of conflict within the immune system. A particularly dangerous locus is a highly variable cluster of NLR genes, DM2, which causes multiple independent incompatibilities with genes that encode a range of biochemical functions, including NLRs. Our findings suggest that deleterious interactions of immune receptors limit the combinations of favorable disease resistance alleles accessible to plant genomes.
Subject(s)
Arabidopsis/genetics , Arabidopsis/immunology , Epistasis, Genetic , Amino Acid Sequence , Arabidopsis/classification , Crosses, Genetic , Genome, Plant , Hybridization, Genetic , Molecular Sequence Data , Phylogeny , Plant Physiological Phenomena , Sequence AlignmentABSTRACT
In this issue of Cell, Chae et al. find that genomic "hot spots" encoding NLR plant immune receptor genes are recurrently responsible for hybrid necrosis, highlighting the role of host-pathogen evolutionary arms races in driving the evolution of hybrid incompatibilities.
Subject(s)
Arabidopsis/genetics , Arabidopsis/immunology , Epistasis, GeneticABSTRACT
Translational reprogramming allows organisms to adapt to changing conditions. Upstream start codons (uAUGs), which are prevalently present in mRNAs, have crucial roles in regulating translation by providing alternative translation start sites1-4. However, what determines this selective initiation of translation between conditions remains unclear. Here, by integrating transcriptome-wide translational and structural analyses during pattern-triggered immunity in Arabidopsis, we found that transcripts with immune-induced translation are enriched with upstream open reading frames (uORFs). Without infection, these uORFs are selectively translated owing to hairpins immediately downstream of uAUGs, presumably by slowing and engaging the scanning preinitiation complex. Modelling using deep learning provides unbiased support for these recognizable double-stranded RNA structures downstream of uAUGs (which we term uAUG-ds) being responsible for the selective translation of uAUGs, and allows the prediction and rational design of translating uAUG-ds. We found that uAUG-ds-mediated regulation can be generalized to human cells. Moreover, uAUG-ds-mediated start-codon selection is dynamically regulated. After immune challenge in plants, induced RNA helicases that are homologous to Ded1p in yeast and DDX3X in humans resolve these structures, allowing ribosomes to bypass uAUGs to translate downstream defence proteins. This study shows that mRNA structures dynamically regulate start-codon selection. The prevalence of this RNA structural feature and the conservation of RNA helicases across kingdoms suggest that mRNA structural remodelling is a general feature of translational reprogramming.
Subject(s)
Codon, Initiator , Nucleic Acid Conformation , RNA, Double-Stranded , RNA, Messenger , Humans , Arabidopsis/genetics , Arabidopsis/immunology , Codon, Initiator/genetics , Innate Immunity Recognition , Open Reading Frames/genetics , Protein Biosynthesis/genetics , Protein Biosynthesis/immunology , Ribosomes/metabolism , RNA, Double-Stranded/chemistry , RNA, Double-Stranded/genetics , RNA, Double-Stranded/metabolism , RNA, Messenger/genetics , Transcriptome , DEAD-box RNA Helicases/chemistry , DEAD-box RNA Helicases/genetics , DEAD-box RNA Helicases/metabolism , Deep LearningABSTRACT
Protein ADP-ribosylation is a reversible post-translational modification that transfers ADP-ribose from NAD+ onto acceptor proteins. Poly(ADP-ribosyl)ation (PARylation), catalyzed by poly(ADP-ribose) polymerases (PARPs) and poly(ADP-ribose) glycohydrolases (PARGs), which remove the modification, regulates diverse cellular processes. However, the chemistry and physiological functions of mono(ADP-ribosyl)ation (MARylation) remain elusive. Here, we report that Arabidopsis zinc finger proteins SZF1 and SZF2, key regulators of immune gene expression, are MARylated by the noncanonical ADP-ribosyltransferase SRO2. Immune elicitation promotes MARylation of SZF1/SZF2 via dissociation from PARG1, which has an unconventional activity in hydrolyzing both poly(ADP-ribose) and mono(ADP-ribose) from acceptor proteins. MARylation antagonizes polyubiquitination of SZF1 mediated by the SH3 domain-containing proteins SH3P1/SH3P2, thereby stabilizing SZF1 proteins. Our study uncovers a noncanonical ADP-ribosyltransferase mediating MARylation of immune regulators and underpins the molecular mechanism of maintaining protein homeostasis by the counter-regulation of ADP-ribosylation and polyubiquitination to ensure proper immune responses.
Subject(s)
ADP-Ribosylation , Arabidopsis Proteins/metabolism , Arabidopsis/immunology , DNA-Binding Proteins/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , Plant Immunity , Ubiquitination , Zinc Fingers , ADP Ribose Transferases/metabolism , Adenosine Diphosphate/chemistry , Arabidopsis/metabolism , CRISPR-Cas Systems , Genes, Plant , Glycoside Hydrolases/metabolism , Homeostasis , Humans , Hydrolysis , Mutation , Plants, Genetically Modified , Poly Adenosine Diphosphate Ribose/metabolism , Poly(ADP-ribose) Polymerases/metabolism , Proteostasis , Seedlings/metabolism , Substrate Specificity , Tristetraprolin/chemistry , Two-Hybrid System Techniques , Ubiquitin/chemistryABSTRACT
Extreme weather conditions associated with climate change affect many aspects of plant and animal life, including the response to infectious diseases. Production of salicylic acid (SA), a central plant defence hormone1-3, is particularly vulnerable to suppression by short periods of hot weather above the normal plant growth temperature range via an unknown mechanism4-7. Here we show that suppression of SA production in Arabidopsis thaliana at 28 °C is independent of PHYTOCHROME B8,9 (phyB) and EARLY FLOWERING 310 (ELF3), which regulate thermo-responsive plant growth and development. Instead, we found that formation of GUANYLATE BINDING PROTEIN-LIKE 3 (GBPL3) defence-activated biomolecular condensates11 (GDACs) was reduced at the higher growth temperature. The altered GDAC formation in vivo is linked to impaired recruitment of GBPL3 and SA-associated Mediator subunits to the promoters of CBP60g and SARD1, which encode master immune transcription factors. Unlike many other SA signalling components, including the SA receptor and biosynthetic genes, optimized CBP60g expression was sufficient to broadly restore SA production, basal immunity and effector-triggered immunity at the elevated growth temperature without significant growth trade-offs. CBP60g family transcription factors are widely conserved in plants12. These results have implications for safeguarding the plant immune system as well as understanding the concept of the plant-pathogen-environment disease triangle and the emergence of new disease epidemics in a warming climate.
Subject(s)
Acclimatization , Arabidopsis Proteins , Arabidopsis , Environment , Global Warming , Plant Immunity , Temperature , Arabidopsis/growth & development , Arabidopsis/immunology , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Calmodulin-Binding Proteins/genetics , Gene Expression Regulation, Plant , Global Warming/statistics & numerical data , Host-Pathogen Interactions , Phytochrome B , Plant Diseases/genetics , Plant Immunity/genetics , Salicylic Acid/metabolism , Transcription FactorsABSTRACT
Plant intracellular nucleotide-binding leucine-rich repeat receptors (NLRs) detect pathogen effectors to trigger immune responses1. Indirect recognition of a pathogen effector by the dicotyledonous Arabidopsis thaliana coiled-coil domain containing NLR (CNL) ZAR1 induces the formation of a large hetero-oligomeric protein complex, termed the ZAR1 resistosome, which functions as a calcium channel required for ZAR1-mediated immunity2-4. Whether the resistosome and channel activities are conserved among plant CNLs remains unknown. Here we report the cryo-electron microscopy structure of the wheat CNL Sr355 in complex with the effector AvrSr356 of the wheat stem rust pathogen. Direct effector binding to the leucine-rich repeats of Sr35 results in the formation of a pentameric Sr35-AvrSr35 complex, which we term the Sr35 resistosome. Wheat Sr35 and Arabidopsis ZAR1 resistosomes bear striking structural similarities, including an arginine cluster in the leucine-rich repeats domain not previously recognized as conserved, which co-occurs and forms intramolecular interactions with the 'EDVID' motif in the coiled-coil domain. Electrophysiological measurements show that the Sr35 resistosome exhibits non-selective cation channel activity. These structural insights allowed us to generate new variants of closely related wheat and barley orphan NLRs that recognize AvrSr35. Our data support the evolutionary conservation of CNL resistosomes in plants and demonstrate proof of principle for structure-based engineering of NLRs for crop improvement.
Subject(s)
Calcium Channels , Cryoelectron Microscopy , NLR Proteins , Plant Proteins , Receptors, Immunologic , Triticum , Arabidopsis/immunology , Arabidopsis/metabolism , Arginine , Calcium Channels/chemistry , Calcium Channels/immunology , Calcium Channels/metabolism , Cations/metabolism , Leucine , NLR Proteins/chemistry , NLR Proteins/immunology , NLR Proteins/metabolism , Plant Diseases/immunology , Plant Diseases/microbiology , Plant Immunity , Plant Proteins/chemistry , Plant Proteins/immunology , Plant Proteins/metabolism , Receptors, Immunologic/chemistry , Receptors, Immunologic/immunology , Receptors, Immunologic/metabolism , Triticum/immunology , Triticum/metabolism , Amino Acid Motifs , Conserved Sequence , ElectrophysiologyABSTRACT
The sensing of microbe-associated molecular patterns (MAMPs) triggers innate immunity in animals and plants. Lipopolysaccharide (LPS) from Gram-negative bacteria is a potent MAMP for mammals, with the lipid A moiety activating proinflammatory responses via Toll-like receptor 4 (TLR4). Here we found that the plant Arabidopsis thaliana specifically sensed LPS of Pseudomonas and Xanthomonas. We isolated LPS-insensitive mutants defective in the bulb-type lectin S-domain-1 receptor-like kinase LORE (SD1-29), which were hypersusceptible to infection with Pseudomonas syringae. Targeted chemical degradation of LPS from Pseudomonas species suggested that LORE detected mainly the lipid A moiety of LPS. LORE conferred sensitivity to LPS onto tobacco after transient expression, which demonstrated a key function in LPS sensing and indicated the possibility of engineering resistance to bacteria in crop species.
Subject(s)
Arabidopsis Proteins/immunology , Arabidopsis/immunology , Gene Expression Regulation, Plant/immunology , Plant Immunity/genetics , Protein Serine-Threonine Kinases/immunology , Arabidopsis/drug effects , Arabidopsis/genetics , Arabidopsis/microbiology , Arabidopsis Proteins/genetics , Lipopolysaccharides/pharmacology , Protein Serine-Threonine Kinases/genetics , Pseudomonas syringae/chemistry , Pseudomonas syringae/immunology , Signal Transduction , Nicotiana/drug effects , Nicotiana/genetics , Nicotiana/immunology , Nicotiana/microbiology , Transgenes , Xanthomonas campestris/chemistry , Xanthomonas campestris/immunologyABSTRACT
Calcium-dependent protein kinases (CPKs) can decode and translate intracellular calcium signals to induce plant immunity. Mutation of the exocyst subunit gene EXO70B1 causes autoimmunity that depends on CPK5 and the Toll/interleukin-1 receptor (TIR) domain resistance protein TIR-NBS2 (TN2), where direct interaction with TN2 stabilizes CPK5 kinase activity. However, how the CPK5-TN2 interaction initiates downstream immune responses remains unclear. Here, we show that, besides CPK5 activity, the physical interaction between CPK5 and functional TN2 triggers immune activation in exo70B1 and may represent reciprocal regulation between CPK5 and the TIR domain functions of TN2 in Arabidopsis (Arabidopsis thaliana). Moreover, we detected differential phosphorylation of the calmodulin-binding transcription activator 3 (CAMTA3) in the cpk5 background. CPK5 directly phosphorylates CAMTA3 at S964, contributing to its destabilization. The gain-of-function CAMTA3A855V variant that resists CPK5-induced degradation rescues immunity activated through CPK5 overexpression or exo70B1 mutation. Thus, CPK5-mediated immunity is executed through CAMTA3 repressor degradation via phosphorylation-induced and/or calmodulin-regulated processes. Conversely, autoimmunity in camta3 also partially requires functional CPK5. While the TIR domain activity of TN2 remains to be tested, our study uncovers a TN2-CPK5-CAMTA3 signaling module for exo70B1-mediated autoimmunity, highlighting the direct embedding of a calcium-sensing decoder element within resistance signalosomes.