RESUMO
Plants and animals detect biomolecules termed microbe-associated molecular patterns (MAMPs) and induce immunity. Agricultural production is severely impacted by pathogens which can be controlled by transferring immune receptors. However, most studies use a single MAMP epitope and the impact of diverse multicopy MAMPs on immune induction is unknown. Here, we characterized the epitope landscape from five proteinaceous MAMPs across 4,228 plant-associated bacterial genomes. Despite the diversity sampled, natural variation was constrained and experimentally testable. Immune perception in both Arabidopsis and tomato depended on both epitope sequence and copy number variation. For example, Elongation Factor Tu is predominantly single copy, and 92% of its epitopes are immunogenic. Conversely, 99.9% of bacterial genomes contain multiple cold shock proteins, and 46% carry a nonimmunogenic form. We uncovered a mechanism for immune evasion, intrabacterial antagonism, where a nonimmunogenic cold shock protein blocks perception of immunogenic forms encoded in the same genome. These data will lay the foundation for immune receptor deployment and engineering based on natural variation.
Assuntos
Arabidopsis , Epitopos , Solanum lycopersicum , Epitopos/imunologia , Solanum lycopersicum/imunologia , Solanum lycopersicum/genética , Solanum lycopersicum/microbiologia , Arabidopsis/imunologia , Arabidopsis/genética , Genoma Bacteriano , Moléculas com Motivos Associados a Patógenos/imunologia , Moléculas com Motivos Associados a Patógenos/metabolismo , Imunidade Vegetal/genética , Imunidade Vegetal/imunologia , Fator Tu de Elongação de Peptídeos/genética , Fator Tu de Elongação de Peptídeos/imunologia , Proteínas de Bactérias/imunologia , Proteínas de Bactérias/genética , Bactérias/imunologia , Bactérias/genética , Proteínas e Peptídeos de Choque Frio/genética , Proteínas e Peptídeos de Choque Frio/imunologia , Proteínas e Peptídeos de Choque Frio/metabolismoRESUMO
Lysin motif receptor-like kinases (LysM-RLKs) are involved in the perception of chitooligosaccharides (COs) and related lipochitooligosaccharides (LCOs) in plants. Expansion and divergence of the gene family during evolution have led to various roles in symbiosis and defense. By studying proteins of the LYR-IA subclass of LysM-RLKs of the Poaceae, we show here that they are high-affinity LCO-binding proteins with a lower affinity for COs, consistent with a role in LCO perception to establish arbuscular mycorrhiza (AM). In Papilionoid legumes, whole-genome duplication has resulted in two LYR-IA paralogs, MtLYR1 and MtNFP in Medicago truncatula, with MtNFP playing an essential role in root nodule symbiosis with nitrogen-fixing rhizobia. We show that MtLYR1 has retained the ancestral LCO-binding characteristic and is dispensable for AM. Domain swapping between the three LysMs of MtNFP and MtLYR1 and mutagenesis in MtLYR1 suggest that the MtLYR1 LCO-binding site is on the second LysM and that divergence in MtNFP led to better nodulation, but surprisingly with decreased LCO binding. These results suggest that divergence of the LCO-binding site has been important for the evolution of a role of MtNFP in nodulation with rhizobia.
Assuntos
Medicago truncatula , Micorrizas , Medicago truncatula/genética , Medicago truncatula/metabolismo , Proteínas de Plantas/metabolismo , Micorrizas/metabolismo , Simbiose/genética , Quitina/metabolismoRESUMO
Sweet potato (Ipomoea batatas) is an important tuber crop, but also target of numerous insect pests. Intriguingly, the abundant storage protein in tubers, sporamin, has intrinsic trypsin protease inhibitory activity. In leaves, sporamin is induced by wounding or a volatile homoterpene and enhances insect resistance. While the signalling pathway leading to sporamin synthesis is partially established, the initial event, perception of a stress-related signal is still unknown. Here, we identified an IbLRR-RK1 that is induced upon wounding and herbivory, and related to peptide-elicitor receptors (PEPRs) from tomato and Arabidopsis. We also identified a gene encoding a precursor protein comprising a peptide ligand (IbPep1) for IbLRR-RK1. IbPep1 represents a distinct signal in sweet potato, which might work in a complementary and/or parallel pathway to the previously described hydroxyproline-rich systemin (HypSys) peptides to strengthen insect resistance. Notably, an interfamily compatibility in the Pep/PEPR system from Convolvulaceae and Solanaceae was identified.
Assuntos
Ipomoea batatas , Ipomoea batatas/genética , Ipomoea batatas/metabolismo , Ligantes , Proteínas de Plantas/metabolismo , Regulação da Expressão Gênica de Plantas , Peptídeos/metabolismoRESUMO
Plant -specific lysin-motif receptor-like kinases (LysM-RLKs) are implicated in the perception of N-acetyl glucosamine-containing compounds, some of which are important signal molecules in plant-microbe interactions. Among these, both lipo-chitooligosaccharides (LCOs) and chitooligosaccharides (COs) are proposed as arbuscular mycorrhizal (AM) fungal symbiotic signals. COs can also activate plant defence, although there are scarce data about CO production by pathogens, especially nonfungal pathogens. We tested Medicago truncatula mutants in the LysM-RLK MtLYK9 for their abilities to interact with the AM fungus Rhizophagus irregularis and the oomycete pathogen Aphanomyces euteiches. This prompted us to analyse whether A. euteiches can produce COs. Compared with wild-type plants, Mtlyk9 mutants had fewer infection events and were less colonised by the AM fungus. By contrast, Mtlyk9 mutants were more heavily infected by A. euteiches and showed more disease symptoms. Aphanomyces euteiches was also shown to produce short COs, mainly CO II, but also CO III and CO IV, and traces of CO V, both ex planta and in planta. MtLYK9 thus has a dual role in plant immunity and the AM symbiosis, which raises questions about the functioning and the ancestral origins of such a receptor protein.
Assuntos
Glomeromycota/fisiologia , Medicago truncatula/microbiologia , Micorrizas/fisiologia , Imunidade Vegetal , Proteínas de Plantas/metabolismo , Simbiose , Sequência de Aminoácidos , Aphanomyces/fisiologia , Quitina/análogos & derivados , Quitina/biossíntese , Quitosana , Regulação da Expressão Gênica de Plantas , Medicago truncatula/genética , Mutação/genética , Oligossacarídeos , Proteínas de Plantas/química , Proteínas de Plantas/genéticaRESUMO
LYR3 [LysM (lysin motif) receptor-like kinase 3] of Medicago truncatula is a high-affinity binding protein for symbiotic LCO (lipo-chitooligosaccharide) signals, produced by rhizobia bacteria and arbuscular mycorrhizal fungi. The present study shows that LYR3 from several other legumes, but not from two Lupinus species which are incapable of forming the mycorrhizal symbiosis, bind LCOs with high affinity and discriminate them from COs (chitooligosaccharides). The biodiversity of these proteins and the lack of binding to the Lupinus proteins were used to identify features required for high-affinity LCO binding. Swapping experiments between each of the three LysMs of the extracellular domain of the M. truncatula and Lupinus angustifolius LYR3 proteins revealed the crucial role of the third LysM in LCO binding. Site-directed mutagenesis identified a tyrosine residue, highly conserved in all LYR3 LCO-binding proteins, which is essential for high-affinity binding. Molecular modelling suggests that it may be part of a hydrophobic tunnel able to accommodate the LCO acyl chain. The lack of conservation of these features in the binding site of plant LysM proteins binding COs provides a mechanistic explanation of how LCO recognition might differ from CO perception by structurally related LysM receptors.
Assuntos
Quitina/análogos & derivados , Medicago truncatula/metabolismo , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Quitina/metabolismo , Quitosana , Lupinus/metabolismo , Oligossacarídeos , Proteínas de Plantas/genética , Ligação Proteica , Transdução de Sinais , Simbiose/genética , Simbiose/fisiologiaRESUMO
Lipo-chitooligosaccharides produced by nitrogen-fixing rhizobia are signaling molecules involved in the establishment of an important agronomical and ecological symbiosis with plants. These compounds, known as Nod factors, are biologically active on plant roots at very low concentrations indicating that they are perceived by specific receptors. This article summarizes the main strategies developed for the syntheses of bioactive Nod factors and their derivatives in order to better understand their mode of perception. Different Nod factor receptors and LCO-binding proteins identified by genetic or biochemical approaches are also presented, indicating perception mechanisms that seem to be more complicated than expected, probably involving multi-component receptor complexes.
Assuntos
Lipopolissacarídeos/metabolismo , Fixação de Nitrogênio/genética , Rhizobiaceae/metabolismo , Simbiose/genética , Fabaceae/metabolismo , Fabaceae/microbiologia , Lipopolissacarídeos/química , Lipopolissacarídeos/genética , Nitrogênio/metabolismo , Filogenia , Raízes de Plantas/metabolismo , Raízes de Plantas/microbiologia , Rhizobiaceae/genética , Transdução de SinaisRESUMO
Recognition of microbial patterns by host pattern recognition receptors is a key step in immune activation in multicellular eukaryotes. Peptidoglycans (PGNs) are major components of bacterial cell walls that possess immunity-stimulating activities in metazoans and plants. Here we show that PGN sensing and immunity to bacterial infection in Arabidopsis thaliana requires three lysin-motif (LysM) domain proteins. LYM1 and LYM3 are plasma membrane proteins that physically interact with PGNs and mediate Arabidopsis sensitivity to structurally different PGNs from gram-negative and gram-positive bacteria. lym1 and lym3 mutants lack PGN-induced changes in transcriptome activity patterns, but respond to fungus-derived chitin, a pattern structurally related to PGNs, in a wild-type manner. Notably, lym1, lym3, and lym3 lym1 mutant genotypes exhibit supersusceptibility to infection with virulent Pseudomonas syringae pathovar tomato DC3000. Defects in basal immunity in lym3 lym1 double mutants resemble those observed in lym1 and lym3 single mutants, suggesting that both proteins are part of the same recognition system. We further show that deletion of CERK1, a LysM receptor kinase that had previously been implicated in chitin perception and immunity to fungal infection in Arabidopsis, phenocopies defects observed in lym1 and lym3 mutants, such as peptidoglycan insensitivity and enhanced susceptibility to bacterial infection. Altogether, our findings suggest that plants share with metazoans the ability to recognize bacterial PGNs. However, as Arabidopsis LysM domain proteins LYM1, LYM3, and CERK1 form a PGN recognition system that is unrelated to metazoan PGN receptors, we propose that lineage-specific PGN perception systems have arisen through convergent evolution.
Assuntos
Proteínas de Arabidopsis/metabolismo , Bactérias/metabolismo , Peptidoglicano/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/microbiologia , Proteínas de Arabidopsis/classificação , Proteínas de Arabidopsis/genética , Bactérias/crescimento & desenvolvimento , Bactérias/imunologia , Resistência à Doença/genética , Resistência à Doença/imunologia , Regulação da Expressão Gênica de Plantas , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Immunoblotting , Microscopia Confocal , Mutação , Análise de Sequência com Séries de Oligonucleotídeos , Peptidoglicano/imunologia , Filogenia , Doenças das Plantas/genética , Doenças das Plantas/imunologia , Doenças das Plantas/microbiologia , Plantas Geneticamente Modificadas , Proteínas Serina-Treonina Quinases/genética , Pseudomonas syringae/imunologia , Pseudomonas syringae/metabolismo , Pseudomonas syringae/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Staphylococcus aureus/imunologia , Staphylococcus aureus/metabolismo , Staphylococcus aureus/fisiologia , TranscriptomaRESUMO
Plants and animals detect biomolecules termed Microbe-Associated Molecular Patterns (MAMPs) and induce immunity. Agricultural production is severely impacted by pathogens which can be controlled by transferring immune receptors. However, most studies use a single MAMP epitope and the impact of diverse multi-copy MAMPs on immune induction is unknown. Here we characterized the epitope landscape from five proteinaceous MAMPs across 4,228 plant-associated bacterial genomes. Despite the diversity sampled, natural variation was constrained and experimentally testable. Immune perception in both Arabidopsis and tomato depended on both epitope sequence and copy number variation. For example, Elongation Factor Tu is predominantly single copy and 92% of its epitopes are immunogenic. Conversely, 99.9% of bacterial genomes contain multiple Cold Shock Proteins and 46% carry a non-immunogenic form. We uncovered a new mechanism for immune evasion, intrabacterial antagonism, where a non-immunogenic Cold Shock Protein blocks perception of immunogenic forms encoded in the same genome. These data will lay the foundation for immune receptor deployment and engineering based on natural variation.
RESUMO
Plants use surface resident and intracellular immune receptors to provide robust immunity against microbial infections. The contribution of the two receptor types to plant immunity differs spatially and temporally. The ongoing identification of new plant cell surface immune receptors and their microbial-derived immunogenic ligands reveal a previously unexpected complexity of plant surface sensors involved in the detection of specific microbial species. Comparative analyses of the plant species distribution of cell surface immune receptors indicate that plants harbor larger sets of genus- or species-specific surface receptors in addition to very few widespread pattern sensors. Leucine-rich repeat surface and intracellular immune sensors emerge as two polymorphic receptor classes whose evolutionary trajectories appear to be linked. This is consistent with their functional cooperativity in providing full plant immunity.
Assuntos
Células Vegetais , Plantas , Plantas/genética , Imunidade Vegetal/genética , Doenças das PlantasRESUMO
Cell wall integrity (CWI) maintenance is central for plant cells. Mechanical and chemical distortions, pH changes, and breakdown products of cell wall polysaccharides activate plasma membrane-localized receptors and induce appropriate downstream responses. Microbial interactions alter or destroy the structure of the plant cell wall, connecting CWI maintenance to immune responses. Cellulose is the major polysaccharide in the primary and secondary cell wall. Its breakdown generates short-chain cellooligomers that induce Ca2+-dependent CWI responses. We show that these responses require the malectin domain-containing CELLOOLIGOMER-RECEPTOR KINASE 1 (CORK1) in Arabidopsis and are preferentially activated by cellotriose (CT). CORK1 is required for cellooligomer-induced cytoplasmic Ca2+ elevation, reactive oxygen species (ROS) production, mitogen-associated protein kinase (MAPK) activation, cellulose synthase phosphorylation, and the regulation of CWI-related genes, including those involved in biosynthesis of cell wall material, secondary metabolites and tryptophan. Phosphoproteome analyses identified early targets involved in signaling, cellulose synthesis, the endoplasmic reticulum/Golgi secretory pathway, cell wall repair and immune responses. Two conserved phenylalanine residues in the malectin domain are crucial for CORK1 function. We propose that CORK1 is required for CWI and immune responses activated by cellulose breakdown products.
Assuntos
Arabidopsis , Arabidopsis/metabolismo , Celulose/metabolismo , Mitógenos/metabolismo , Fenilalanina/metabolismo , Proteínas Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Triptofano/metabolismoRESUMO
The geomagnetic field (GMF) is one of the environmental stimuli that plants experience continuously on Earth; however, the actions of the GMF on plants are poorly understood. Here, we carried out a time-course microarray experiment to identify genes that are differentially regulated by the GMF in shoot and roots. We also used qPCR to validate the activity of some genes selected from the microarray analysis in a dose-dependent magnetic field experiment. We found that the GMF regulated genes in both shoot and roots, suggesting that both organs can sense the GMF. However, 49% of the genes were regulated in a reverse direction in these organs, meaning that the resident signaling networks define the up- or downregulation of specific genes. The set of GMF-regulated genes strongly overlapped with various stress-responsive genes, implicating the involvement of one or more common signals, such as reactive oxygen species, in these responses. The biphasic dose response of GMF-responsive genes indicates a hormetic response of plants to the GMF. At present, no evidence exists to indicate any evolutionary advantage of plant adaptation to the GMF; however, plants can sense and respond to the GMF using the signaling networks involved in stress responses.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas/efeitos da radiação , Campos Magnéticos , Raízes de Plantas/metabolismo , Brotos de Planta/metabolismo , Transcriptoma/efeitos da radiação , Arabidopsis/genética , Arabidopsis/efeitos da radiação , Proteínas de Arabidopsis/genética , Perfilação da Expressão Gênica , Raízes de Plantas/genética , Raízes de Plantas/efeitos da radiação , Brotos de Planta/genética , Brotos de Planta/efeitos da radiaçãoRESUMO
Bacterial flagella are perceived by the innate immune systems of plants1 and animals2 alike, triggering resistance. Common to higher plants is the immunoreceptor FLAGELLIN-SENSING 2 (FLS2)3, which detects flagellin via its most conserved epitope, flg22. Agrobacterium tumefaciens, which causes crown gall disease in many crop plants, has a highly diverged flg22 epitope and evades immunodetection by plants so far studied. We asked whether, as a next step in this game of 'hide and seek', there are plant species that have evolved immunoreceptors with specificity for the camouflaged flg22Atum of A. tumefaciens. In the wild grape species Vitis riparia, we discovered FLS2XL, a previously unknown form of FLS2, that provides exquisite sensitivity to typical flg22 and to flg22Atum. As exemplified by ectopic expression in tobacco, FLS2XL can limit crown gall disease caused by A. tumefaciens.
Assuntos
Agrobacterium tumefaciens/fisiologia , Flagelina/metabolismo , Proteínas de Plantas/metabolismo , Tumores de Planta/microbiologia , Proteínas Quinases/metabolismo , Vitis/metabolismo , Vitis/microbiologiaRESUMO
In plants, antimicrobial immune responses involve the cellular release of anions and are responsible for the closure of stomatal pores. Detection of microbe-associated molecular patterns (MAMPs) by pattern recognition receptors (PRRs) induces currents mediated via slow-type (S-type) anion channels by a yet not understood mechanism. Here, we show that stomatal closure to fungal chitin is conferred by the major PRRs for chitin recognition, LYK5 and CERK1, the receptor-like cytoplasmic kinase PBL27, and the SLAH3 anion channel. PBL27 has the capacity to phosphorylate SLAH3, of which S127 and S189 are required to activate SLAH3. Full activation of the channel entails CERK1, depending on PBL27. Importantly, both S127 and S189 residues of SLAH3 are required for chitin-induced stomatal closure and anti-fungal immunity at the whole leaf level. Our results demonstrate a short signal transduction module from MAMP recognition to anion channel activation, and independent of ABA-induced SLAH3 activation.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/imunologia , Regulação da Expressão Gênica de Plantas , Canais Iônicos/metabolismo , Estômatos de Plantas/fisiologia , Proteínas Quinases/metabolismo , Arabidopsis/efeitos dos fármacos , Quitina/imunologia , Fungos/química , Estômatos de Plantas/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Reconhecimento de Padrão/metabolismoRESUMO
The Earth magnetic field (or geomagnetic field, GMF) is a natural component of our planet and variations of the GMF are perceived by plants with a still uncharacterized magnetoreceptor. The purpose of this work was to assess the effect of near null magnetic field (NNMF, â¼40 nT) on Arabidopsis thaliana Col0 root ion modulation. A time-course (from 10â¯min to 96â¯h) exposure of Arabidopsis to NNMF was compared to GMF and the content of some cations (NH4+, K+, Ca2+ and Mg2+) and anions (Cl-, SO4=, NO3- and PO4=) was evaluated by capillary electrophoresis. The expression of several cation and anion channel- and transporter-related genes was assessed by gene microarray. A few minutes after exposure to NNMF, Arabidopsis roots responded with a significant change in the content and gene expression of all nutrient ions under study, indicating the presence of a plant magnetoreceptor that responds immediately to MF variations by modulating channels, transporters and genes involved in mineral nutrition. The response of Arabidopsis to reduced MF was a general reduction of plant ion uptake and transport. Our data suggest the importance to understand the nature and function of the plant magnetoreceptor for future space programs involving plant growth in environments with a reduced MF.
Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Campos Magnéticos , Minerais/metabolismo , Raízes de Plantas/metabolismo , Arabidopsis/genética , Arabidopsis/efeitos da radiação , Perfilação da Expressão Gênica , Raízes de Plantas/genética , Raízes de Plantas/efeitos da radiaçãoRESUMO
The discovery in tomato of systemin, the first plant peptide hormone1,2, was a fundamental change for the concept of plant hormones. Numerous other peptides have since been shown to play regulatory roles in many aspects of the plant life, including growth, development, fertilization and interactions with symbiotic organisms3-6. Systemin, an 18 amino acid peptide derived from a larger precursor protein 7 , was proposed to act as the spreading signal that triggers systemic defence responses observed in plants after wounding or attack by herbivores1,7,8. Further work culminated in the identification of a leucine-rich repeat receptor kinase (LRR-RK) as the systemin receptor 160 (SR160)9,10. SR160 is a tomato homologue of Brassinosteroid Insensitive 1 (BRI1), which mediates the regulation of growth and development in response to the steroid hormone brassinolide11-13. However, a role of SR160/BRI1 as systemin receptor could not be corroborated by others14-16. Here, we demonstrate that perception of systemin depends on a pair of distinct LRR-RKs termed SYR1 and SYR2. SYR1 acts as a genuine systemin receptor that binds systemin with high affinity and specificity. Further, we show that presence of SYR1, although not decisive for local and systemic wound responses, is important for defence against insect herbivory.
Assuntos
Peptídeos/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Proteínas de Plantas/metabolismo , Receptores de Superfície Celular/metabolismo , Solanum lycopersicum/metabolismo , Herbivoria , Especificidade por SubstratoRESUMO
The Avr1b locus is required for avirulence of the oomycete pathogen Phytophthora sojae on soybeans carrying resistance gene Rps1b. One of the Avr genes of the locus (Avr1b-1) was shown to encode an elicitor. We have analyzed the spatial and temporal expression patterns of Avr1b-1 in comparison to defense-related genes induced in soybean. Avr1b-1 expression was detectable mainly in close proximity to the site of infection, in wound-inoculated hypocotyls as well as in roots infected with zoospores. Usually, in compatible interactions, higher expression levels of Avr1b-1 were observed in roots when compared with incompatible P. sojae-soybean interactions, whereas neither the timing nor the amount of transcript accumulation of defense-related genes showed cultivar-specific differences. In contrast, the PsojNIP gene encoding a proposed virulence factor was expressed only during the necrotrophic phase in the compatible interaction.
Assuntos
Glycine max/genética , Phytophthora/genética , Phytophthora/patogenicidade , Doenças das Plantas/genética , Actinas/biossíntese , Actinas/genética , Expressão Gênica , Hipocótilo/crescimento & desenvolvimento , Micélio/crescimento & desenvolvimento , Phytophthora/crescimento & desenvolvimento , Phytophthora/metabolismo , Doenças das Plantas/parasitologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Glycine max/metabolismo , Glycine max/parasitologia , Tubulina (Proteína)/biossíntese , Tubulina (Proteína)/genéticaRESUMO
LYR3, LYK3, and NFP are lysin motif-containing receptor-like kinases (LysM-RLKs) from Medicago truncatula, involved in perception of symbiotic lipo-chitooligosaccharide (LCO) signals. Here, we show that LYR3, a high-affinity LCO-binding protein, physically interacts with LYK3, a key player regulating symbiotic interactions. In vitro, LYR3 is phosphorylated by the active kinase domain of LYK3. Fluorescence lifetime imaging/Förster resonance energy transfer (FLIM/FRET) experiments in tobacco protoplasts show that the interaction between LYR3 and LYK3 at the plasma membrane is disrupted or inhibited by addition of LCOs. Moreover, LYR3 attenuates the cell death response, provoked by coexpression of NFP and LYK3 in tobacco leaves.
Assuntos
Lipopolissacarídeos/metabolismo , Medicago truncatula/metabolismo , Proteínas Quinases/metabolismo , Protoplastos/metabolismo , Membrana Celular/metabolismo , Transferência Ressonante de Energia de Fluorescência , Medicago truncatula/química , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Ligação Proteica , Proteínas Quinases/química , Proteínas Quinases/genética , Simbiose , Nicotiana/genética , Nicotiana/metabolismoRESUMO
The beta-glucan-binding protein (GBP) of soybean (Glycine max L.) has been shown to contain two different activities. As part of the plasma membrane-localized pathogen receptor complex, it binds a microbial cell wall elicitor, triggering the activation of defence responses. Additionally, the GBP is able to hydrolyze beta-1,3-glucans, as present in the cell walls of potential pathogens. The substrate specificity, the mode of action, and the stereochemistry of the catalysis have been elucidated. This defines for the first time the inverting mode of the catalytic mechanism of glycoside hydrolases belonging to family 81.
Assuntos
Proteínas de Transporte/metabolismo , Glycine max/química , Lectinas/metabolismo , Calmodulina/metabolismo , Proteínas de Ligação a Calmodulina/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/isolamento & purificação , Catálise , Clonagem Molecular , Glucanos/metabolismo , Glicosídeo Hidrolases , Hidrólise , Lectinas/genética , Lectinas/isolamento & purificação , Proteínas de Plantas , Estereoisomerismo , Especificidade por SubstratoRESUMO
N-acetylglucosamine-based saccharides (chitosaccharides) are components of microbial cell walls and act as molecular signals during host-microbe interactions. In the legume plant Medicago truncatula, the perception of lipochitooligosaccharide signals produced by symbiotic rhizobia and arbuscular mycorrhizal fungi involves the Nod Factor Perception (NFP) lysin motif receptor-like protein and leads to the activation of the so-called common symbiotic pathway. In rice and Arabidopsis, lysin motif receptors are involved in the perception of chitooligosaccharides released by pathogenic fungi, resulting in the activation of plant immunity. Here we report the structural characterization of atypical chitosaccharides from the oomycete pathogen Aphanomyces euteiches, and their biological activity on the host Medicago truncatula. Using a combination of biochemical and biophysical approaches, we show that these chitosaccharides are linked to ß-1,6-glucans, and contain a ß-(1,3;1,4)-glucan backbone whose ß-1,3-linked glucose units are substituted on their C-6 carbon by either glucose or N-acetylglucosamine residues. This is the first description of this type of structural motif in eukaryotic cell walls. Glucan-chitosaccharide fractions of A. euteiches induced the expression of defense marker genes in Medicago truncatula seedlings independently from the presence of a functional Nod Factor Perception protein. Furthermore, one of the glucan-chitosaccharide fractions elicited calcium oscillations in the nucleus of root cells. In contrast to the asymmetric oscillatory calcium spiking induced by symbiotic lipochitooligosaccharides, this response depends neither on the Nod Factor Perception protein nor on the common symbiotic pathway. These findings open new perspectives in oomycete cell wall biology and elicitor recognition and signaling in legumes.
Assuntos
Aphanomyces/citologia , Sinalização do Cálcio/efeitos dos fármacos , Parede Celular/química , Quitina/farmacologia , Glucanos/farmacologia , Medicago truncatula/genética , Medicago truncatula/imunologia , Acetilglucosamina/metabolismo , Sinalização do Cálcio/genética , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Quitina/química , Cromatografia em Gel , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genes de Plantas , Glucanos/química , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Medicago truncatula/microbiologia , Modelos Moleculares , Epiderme Vegetal/citologia , Epiderme Vegetal/efeitos dos fármacos , Raízes de Plantas/citologia , Raízes de Plantas/efeitos dos fármacosRESUMO
While chitooligosaccharides (COs) derived from fungal chitin are potent elicitors of defense reactions, structurally related signals produced by certain bacteria and fungi, called lipo-chitooligosaccharides (LCOs), play important roles in the establishment of symbioses with plants. Understanding how plants distinguish between friend and foe through the perception of these signals is a major challenge. We report the synthesis of a range of COs and LCOs, including photoactivatable probes, to characterize a membrane protein from the legume Medicago truncatula. By coupling photoaffinity labeling experiments with proteomics and transcriptomics, we identified the likely LCO-binding protein as LYR3, a lysin motif receptor-like kinase (LysM-RLK). LYR3, expressed heterologously, exhibits high-affinity binding to LCOs but not COs. Homology modeling, based on the Arabidopsis CO-binding LysM-RLK AtCERK1, suggests that LYR3 could accommodate the LCO in a conserved binding site. The identification of LYR3 opens up ways for the molecular characterization of LCO/CO discrimination.