RESUMEN
The Schizosaccharomyces pombe Clr6S complex, a class I histone deacetylase complex, functions as a zinc-dependent enzyme to remove acetyl groups from lysine residues in histone tails. We report here the cryo-EM structure of Clr6S alone and a cryo-EM map of Clr6S in complex with a nucleosome. The active center, revealed at near-atomic resolution, includes features important for catalysis-A water molecule coordinated by zinc, the likely nucleophile for attack on the acetyl-lysine bond, and a loop that may position the substrate for catalysis. The cryo-EM map in the presence of a nucleosome reveals multiple Clr6S-nucleosome contacts and a high degree of relative motion of Clr6S and the nucleosome. Such flexibility may be attributed to interaction at a site in the flexible histone tail and is likely important for the function of the deacetylase, which acts at multiple sites in other histone tails.
Asunto(s)
Histonas , Schizosaccharomyces , Histonas/genética , Nucleosomas , Lisina/química , Histona Desacetilasas/metabolismo , Schizosaccharomyces/metabolismo , ZincRESUMEN
Nucleosome acetyltransferase of H4 (NuA4), one of two major histone acetyltransferase complexes in Saccharomyces cerevisiae specifically acetylates histone H2A and H4, resulting in increased transcriptional activity. Here we present a 3.8-4.0 Å resolution structure of the NuA4 complex from cryoelectron microscopy and associated biochemical studies. The determined structure comprises six subunits and appropriately 5,000 amino acids, with a backbone formed by subunits Eaf1 and Eaf2 spanning from an Actin-Arp4 module to a platform subunit Tra1. Seven subunits are missing from the cryo-EM map. The locations of missing components, Yaf9, and three subunits of the Piccolo module Esa1, Yng2, and Eaf6 were determined. Biochemical studies showed that the Piccolo module and the complete NuA4 exhibit comparable histone acetyltransferase activities, but the Piccolo module binds to nucleosomes, whereas the complete NuA4 does not. The interaction lifetime of NuA4 and nucleosome is evidently short, possibly because of subunits of the NuA4 complex that diminish the affinity of the Piccolo module for the nucleosome, enabling rapid movement from nucleosome to nucleosome.
Asunto(s)
Nucleosomas , Proteínas de Saccharomyces cerevisiae , Nucleosomas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Histona Acetiltransferasas/metabolismo , Microscopía por Crioelectrón , Saccharomyces cerevisiae/metabolismo , Histonas/metabolismoRESUMEN
The oncogenic fusion protein BCR-ABL is the driving force of leukemogenesis in chronic myeloid leukemia (CML). Despite the great advance in CML treatment through the application of tyrosine kinase inhibitors (TKIs) against BCR-ABL, disease recurrence after TKI discontinuation and clinical resistance mainly due to BCR-ABL mutations continue to be an issue. Herein we report our efforts to synthesize a novel series of CRBN-recruiting proteolysis-targeting chimeras (PROTACs) targeting BCR-ABL based on the allosteric inhibitor asciminib. Our efforts have led to the discovery of compound 30 (SIAIS100) through extensive SAR studies by the optimization of linker parameters as well as linker attachment points of both target-binding warhead and CRBN ligands, which exhibited the most potent degradative activity with a DC50 value of 2.7 nM and Dmax of 91.2% against BCR-ABL and has an IC50 value of 12 nM in BCR-ABL + K562 cells. The binding model and the stability evaluation of 30-induced ternary complex formation were also elucidated through computational simulations. Furthermore, 30 induced sustained and robust BCR-ABL degradation and maintained the efficacy for 96 h post-washout. Moreover, the proteomics analysis showed that 30 degraded BCR-ABL and three CRBN's neo-substrates, including IKZF1, IKZF3, and ZFP91. Additionally, 30 also exerted degradative activity against a panel of clinically relevant resistance-conferring mutations of BCR-ABL, including gatekeeper mutation T315I, several single mutations associated with TKI resistance, and certain highly resistant compound mutations. Our study provided a deeper understanding of the development of PROTACs targeting BCR-ABL and novel potential therapeutic agents for CML treatment.
Asunto(s)
Leucemia Mielógena Crónica BCR-ABL Positiva , Inhibidores de Proteínas Quinasas , Humanos , Inhibidores de Proteínas Quinasas/química , Resistencia a Antineoplásicos , Proteínas de Fusión bcr-abl , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Células K562 , Mutación , Ubiquitina-Proteína LigasasRESUMEN
Mediator is a universal adaptor for transcription control. It serves as an interface between gene-specific activator or repressor proteins and the general RNA polymerase II (pol II) transcription machinery. Previous structural studies revealed a relatively small part of Mediator and none of the gene activator-binding regions. We have determined the cryo-EM structure of the Mediator at near-atomic resolution. The structure reveals almost all amino acid residues in ordered regions, including the major targets of activator proteins, the Tail module, and the Med1 subunit of the Middle module. Comparison of Mediator structures with and without pol II reveals conformational changes that propagate across the entire Mediator, from Head to Tail, coupling activator- and pol II-interacting regions.
Asunto(s)
Subunidad 1 del Complejo Mediador/metabolismo , Aminoácidos/genética , Conformación Proteica , ARN Polimerasa II/metabolismo , Factores de Transcripción/metabolismo , Transcripción Genética/genéticaRESUMEN
Ice-binding proteins (IBPs) are found in many biological kingdoms where they protect organisms from freezing damage as antifreeze agents or inhibitors of ice recrystallization. Here, the crystal structure of recombinant IBP from carrot (Daucus carota) has been solved to a resolution of 2.3â Å. As predicted, the protein is a structural homologue of a plant polygalacturonase-inhibiting protein forming a curved solenoid structure with a leucine-rich repeat motif. Unexpectedly, close examination of its surface did not reveal any large regions of flat, regularly spaced hydrophobic residues that characterize the ice-binding sites (IBSs) of potent antifreeze proteins from freeze-resistant fish and insects. An IBS was defined by site-directed mutagenesis of residues on the convex surface of the carrot solenoid. This imperfect site is reminiscent of the irregular IBS of grass 'antifreeze' protein. Like the grass protein, the carrot IBP has weak freezing point depression activity but is extremely active at nanomolar concentrations in inhibiting ice recrystallization. Ice crystals formed in the presence of both plant proteins grow slowly and evenly in all directions. We suggest that this slow, controlled ice growth is desirable for freeze tolerance. The fact that two plant IBPs have evolved very different protein structures to affect ice in a similar manner suggests this pattern of weak freezing point depression and strong ice recrystallization inhibition helps their host to tolerate freezing rather than to resist it.
Asunto(s)
Proteínas Anticongelantes/química , Proteínas Anticongelantes/metabolismo , Daucus carota/metabolismo , Hielo , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Sitios de Unión , Cristalización , Congelación , Interacciones Hidrofóbicas e Hidrofílicas , Conformación Proteica , Dominios ProteicosRESUMEN
Histidine biosynthesis, which is absent in animals, was shown to be highly conserved among gram-negative bacteria, thus making it an attractive target for antibiotic design. There are many fusion forms of enzymes in the histidine biosynthetic pathway and people still have limited knowledge about their domain organizations and catalytic mechanisms, due to the lack of structural information. Here we report the first crystal structure of Shigella flexneri bi-functional enzyme HisIE (SfHisIE) that functions in the 2nd and 3rd steps in the histidine biosynthetic pathway. This structure shows that HisIE exists as dimers with two loops (fusion loop) connecting the individual dimer of HisE and HisI in its N-terminus and C-terminus respectively. Our mutagenesis study shows mutations in this fusion loop are lethal for bacteria indicating the advantage of gene fusion in Histidine biosynthesis. Structural analysis revealed several highly conserved residues in the putative ligand binding grooves of HisE and HisI, showing an evolutionarily conserved catalytic mechanism shared among gram negative-bacteria.
Asunto(s)
Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Histidina/biosíntesis , Shigella flexneri/enzimología , Secuencia de Aminoácidos , Biocatálisis , Modelos Moleculares , Dominios Proteicos , Estructura Secundaria de ProteínaRESUMEN
Leucine-rich repeat receptor-like kinases (LRR-RLKs) are widespread in different plant species and play important roles in growth and development. Germination inhibition is vital for the completion of seed maturation and cell expansion is a fundamental cellular process driving plant growth. Here, we report genetic and structural characterizations of a functionally uncharacterized LRR-RLK, named GRACE (Germination Repression and Cell Expansion receptor-like kinase). Overexpression of GRACE in Arabidopsis exhibited delayed germination, enlarged cotyledons, rosette leaves and stubbier petioles. Conversely, these phenotypes were reversed in the T-DNA insertion knock-down mutant grace-1 plants. A crystal structure of the extracellular domain of GRACE (GRACE-LRR) determined at the resolution of 3.0 Å revealed that GRACE-LRR assumed a right-handed super-helical structure with an island domain (ID). Structural comparison showed that structure of the ID in GRACE-LRR is strikingly different from those observed in other LRR-RLKs. This structural observation implies that GRACE might perceive a new ligand for signaling. Collectively, our data support roles of GRACE in repressing seed germination and promoting cell expansion of Arabidopsis, presumably by perception of unknown ligand(s).
RESUMEN
Transportation of the immobile sperms directed by pollen tubes to the ovule-enclosed female gametophytes is important for plant sexual reproduction. The defensin-like (DEFL) cysteine-rich peptides (CRPs) LUREs play an essential role in pollen tube attraction to the ovule, though their receptors still remain controversial. Here we provide several lines of biochemical evidence showing that the extracellular domain of the leucine-rich repeat receptor kinase (LRR-RK) PRK6 from Arabidopsis thaliana directly interacts with AtLURE1 peptides. Structural study reveals that a C-terminal loop of the LRR domain (AtPRK6LRR) is responsible for recognition of AtLURE1.2, mediated by a set of residues largely conserved among PRK6 homologs from Arabidopsis lyrata and Capsella rubella, supported by in vitro mutagenesis and semi-in-vivo pollen tube growth assays. Our study provides evidence showing that PRK6 functions as a receptor of the LURE peptides in A. thaliana and reveals a unique ligand recognition mechanism of LRR-RKs.
Asunto(s)
Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Tubo Polínico/metabolismo , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , Secuencia de Aminoácidos , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Capsella/genética , Capsella/metabolismo , Cristalografía por Rayos X , Genes de Plantas , Ligandos , Modelos Moleculares , Mutagénesis Sitio-Dirigida , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Dominios y Motivos de Interacción de Proteínas , Proteínas Serina-Treonina Quinasas/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Homología de Secuencia de Aminoácido , Electricidad EstáticaAsunto(s)
Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Arabidopsis/inmunología , Inmunidad de la Planta , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , Arabidopsis/genética , Plantas Modificadas Genéticamente , Unión Proteica , Dominios Proteicos , Relación Estructura-ActividadRESUMEN
Self-incompatibility (SI) is a widespread mechanism in flowering plants which prevents self-fertilization and inbreeding. In Brassica, recognition of the highly polymorphic S-locus cysteine-rich protein (SCR; or S-locus protein 11) by the similarly polymorphic S-locus receptor kinase (SRK) dictates the SI specificity. Here, we report the crystal structure of the extracellular domain of SRK9 (eSRK9) in complex with SCR9 from Brassica rapa. SCR9 binding induces eSRK9 homodimerization, forming a 2:2 eSRK:SCR heterotetramer with a shape like the letter "A". Specific recognition of SCR9 is mediated through three hyper-variable (hv) regions of eSRK9. Each SCR9 simultaneously interacts with hvI and one-half of hvII from one eSRK9 monomer and the other half of hvII from the second eSRK9 monomer, playing a major role in mediating SRK9 homodimerization without involving interaction between the two SCR9 molecules. Single mutations of residues critical for the eSRK9-SCR9 interaction disrupt their binding in vitro. Our study rationalizes a body of data on specific recognition of SCR by SRK and provides a structural template for understanding the co-evolution between SRK and SCR.
Asunto(s)
Brassica/metabolismo , Proteínas de Plantas/química , Secuencia de Aminoácidos , Cristalografía por Rayos X , Dimerización , Mutagénesis , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Unión Proteica , Proteínas Quinasas/química , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Estructura Cuaternaria de Proteína , Estructura Terciaria de Proteína , Alineación de SecuenciaRESUMEN
Secreted signaling peptides or peptide hormones play crucial roles in plant growth and development through coordination of cell-cell communication. Perception of peptide hormones in plants generally relies on membrane-localized receptor kinases (RKs). Progress has recently been made in structural elucidation of interactions between posttranslationally modified peptide hormones and RKs. The structural studies suggest conserved receptor binding and activation mechanisms of this type of peptide hormones involving their conserved C-termini. Here, we review these structural data and discuss how the conserved mechanisms can be used to match peptide-RK pairs.
Asunto(s)
Hormonas Peptídicas/química , Hormonas Peptídicas/metabolismo , Reguladores del Crecimiento de las Plantas/química , Reguladores del Crecimiento de las Plantas/metabolismo , Proteínas de Plantas/metabolismo , Secuencia de Aminoácidos , Proteínas de Plantas/químicaRESUMEN
In Arabidopsis, the CLAVATA3/EMBRYO SURROUNDING REGION-RELATED (CLE) peptides play important roles in regulating proliferation and differentiation of plant-specific stem cells. Although receptors of CLEs are reported to be leucine-rich repeat receptor kinases, the mechanisms underlying CLE-induced receptor activation remain largely unknown. Here we show that SOMATIC EMBRYOGENESIS RECEPTOR KINASEs (SERKs) serve as co-receptors in CLE41/TDIF-PXY signaling to regulate plant vascular development. TDIF induces interaction of its receptor PXY with SERKs in vitro and in vivo. Furthermore, the serk1-1 serk2-1 bak1-5 mutant plants are less sensitive to TDIF, phenocopying the pxy mutant with a compromised promotion of procambial cell proliferation. Crystal structure of the PXY-TDIF-SERK2 complex reveals that the last amino acid of TDIF conserved among CLEs and other evolutionary-related peptides is important for the interaction between SERK2 and PXY. Taken together, our current study identifies SERKs as signaling components of the TDIF-PXY pathway and suggests a conserved activation mechanism of CLE receptors.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas Quinasas/metabolismo , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Proliferación Celular/genética , Proliferación Celular/fisiología , Regulación de la Expresión Génica de las Plantas , Oligopéptidos/genética , Oligopéptidos/metabolismo , Proteínas Quinasas/genética , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Xilema/crecimiento & desarrollo , Xilema/metabolismoRESUMEN
Peptide-mediated cell-to-cell signaling has crucial roles in coordination and definition of cellular functions in plants. Peptide-receptor matching is important for understanding the mechanisms underlying peptide-mediated signaling. Here we report the structure-guided identification of root meristem growth factor (RGF) receptors important for plant development. An assay based on a signature ligand recognition motif (Arg-x-Arg) conserved in a subfamily of leucine-rich repeat receptor kinases (LRR-RKs) identified the functionally uncharacterized LRR-RK At4g26540 as a receptor of RGF1 (RGFR1). We further solved the crystal structure of RGF1 in complex with the LRR domain of RGFR1 at a resolution of 2.6 Å, which reveals that the Arg-x-Gly-Gly (RxGG) motif is responsible for specific recognition of the sulfate group of RGF1 by RGFR1. Based on the RxGG motif, we identified additional four RGFRs. Participation of the five RGFRs in RGF-induced signaling is supported by biochemical and genetic data. We also offer evidence showing that SERKs function as co-receptors for RGFs. Taken together, our study identifies RGF receptors and co-receptors that can link RGF signals with their downstream components and provides a proof of principle for structure-based matching of LRR-RKs with their peptide ligands.
Asunto(s)
Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Meristema/crecimiento & desarrollo , Meristema/metabolismo , Hormonas Peptídicas/metabolismo , Receptores de Superficie Celular/metabolismo , Secuencia de Aminoácidos , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Ligandos , Mutación con Pérdida de Función , Modelos Biológicos , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de SeñalRESUMEN
Chitin is the major component of fungal cell wall and serves as a molecular pattern that can be recognized by the receptor OsCEBiP in rice, a lysine motif (LysM) receptor-like protein (RLP), to trigger immune responses. The molecular mechanisms underlying chitin recognition remain elusive. Here we report the crystal structures of the ectodomain of OsCEBiP (OsCEBiP-ECD) in free and chitin-bound forms. The structures reveal that OsCEBiP-ECD contains three tandem LysMs followed by a novel structure fold of cysteine-rich domain. The structures showed that chitin binding induces no striking conformational changes in OsCEBiP. Structural comparison among N-acetylglucosamine (NAG) oligomer-bound LysMs revealed a highly conserved recognition mechanism, which is expected to facilitate study of other LysM-containing proteins for their NAG binding. Modeling study showed that chitin induces OsCEBiP homodimerization in a "sliding mode". Our data provide insights into rice chitin receptor-mediated immunity triggered by fungal cell wall.
Asunto(s)
Quitina/metabolismo , Proteínas de Plantas/química , Receptores de Superficie Celular/química , Acetilglucosamina/química , Acetilglucosamina/metabolismo , Sitios de Unión , Pared Celular/química , Pared Celular/metabolismo , Quitina/química , Hongos/química , Simulación del Acoplamiento Molecular , Oryza/química , Proteínas de Plantas/metabolismo , Unión Proteica , Receptores de Superficie Celular/metabolismoRESUMEN
Plants can achieve amazing lifespans because of their continuous and repetitive formation of new organs by stem cells present within meristems. The balance between proliferation and differentiation of meristem cells is largely regulated by the CLAVATA3/ENDOSPERM SURROUNDING REGION (CLE) peptide hormones. One of the well-characterized CLE peptides, CLE41/TDIF (tracheary elements differentiation inhibitory factor), functions to suppress tracheary element differentiation and promote procambial cell proliferation, playing important roles in vascular development and wood formation. The recognition mechanisms of TDIF or other CLE peptides by their respective receptors, however, remain largely elusive. Here we report the crystal structure of TDIF in complex with its receptor PXY, a leucine-rich repeat receptor kinase (LRR-RK). Our structure reveals that TDIF mainly adopts an "Ω"-like conformation binding to the inner surface of the LRR domain of PXY. Interaction between TDIF and PXY is predominately mediated by the relatively conserved amino acids of TDIF. Structure-based sequence alignment showed that the TDIF-interacting motifs are also conserved among other known CLE receptors. Our data provide a structural template for understanding the recognition mechanism of CLE peptides by their receptors, offering an opportunity for the identification of receptors of other uncharacterized CLE peptides.
Asunto(s)
Proteínas de Arabidopsis/sangre , Proteínas de Arabidopsis/química , Oligopéptidos/sangre , Péptidos/metabolismo , Proteínas Quinasas/química , Receptores de Superficie Celular/metabolismo , Secuencia de Aminoácidos , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Secuencia Conservada , Cristalografía por Rayos X , Ligandos , Modelos Moleculares , Mutagénesis/genética , Oligopéptidos/química , Oligopéptidos/metabolismo , Péptidos/química , Unión Proteica , Dominios ProteicosRESUMEN
Toll-like receptors (TLRs) have crucial roles in innate immunity, functioning as pattern-recognition receptors. TLR13 recognizes a conserved sequence from bacterial 23S rRNA and then triggers an immune response. Here we report the crystal structure of the mouse TLR13 ectodomain bound by a 13-nt single-stranded (ss) RNA derived from 23S rRNA. The ssRNA induces TLR13 dimerization but assumes a stem-loop-like structure that is completely different from that in the bacterial ribosome but nevertheless is crucial for TLR13 recognition. Most of the RNA nucleotides are splayed out to make base-specific contacts with the concave surface of TLR13, and RNA-specific interactions are important to allow TLR13 to distinguish RNA from DNA. Interestingly, a viral-derived 16-nt ssRNA predicted to form a similar stem-loop-like structure also induces TLR13 activation. Together, our results reveal the structural mechanism of TLR13's sequence- and conformation-specific recognition of ssRNA.
Asunto(s)
Modelos Moleculares , ARN Ribosómico 23S/química , ARN Ribosómico 23S/metabolismo , Receptores Toll-Like/química , Receptores Toll-Like/metabolismo , Animales , Cromatografía en Gel , Microscopía por Crioelectrón , Cristalización , Dimerización , Procesamiento de Imagen Asistido por Computador , Luciferasas , Ratones , Microscopía Electrónica de Transmisión , Oligonucleótidos/genética , Unión Proteica , Conformación Proteica , ARN Ribosómico 23S/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores Toll-Like/genéticaRESUMEN
Phytosulfokine (PSK) is a disulfated pentapeptide that has a ubiquitous role in plant growth and development. PSK is perceived by its receptor PSKR, a leucine-rich repeat receptor kinase (LRR-RK). The mechanisms underlying the recognition of PSK, the activation of PSKR and the identity of the components downstream of the initial binding remain elusive. Here we report the crystal structures of the extracellular LRR domain of PSKR in free, PSK- and co-receptor-bound forms. The structures reveal that PSK interacts mainly with a ß-strand from the island domain of PSKR, forming an anti-ß-sheet. The two sulfate moieties of PSK interact directly with PSKR, sensitizing PSKR recognition of PSK. Supported by biochemical, structural and genetic evidence, PSK binding enhances PSKR heterodimerization with the somatic embryogenesis receptor-like kinases (SERKs). However, PSK is not directly involved in PSKR-SERK interaction but stabilizes PSKR island domain for recruitment of a SERK. Our data reveal the structural basis for PSKR recognition of PSK and allosteric activation of PSKR by PSK, opening up new avenues for the design of PSKR-specific small molecules.
Asunto(s)
Proteínas de Arabidopsis/agonistas , Proteínas de Arabidopsis/química , Arabidopsis/química , Reguladores del Crecimiento de las Plantas/química , Reguladores del Crecimiento de las Plantas/farmacología , Receptores de Superficie Celular/agonistas , Receptores de Superficie Celular/química , Regulación Alostérica/efectos de los fármacos , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Cristalografía por Rayos X , Modelos Moleculares , Mutación/genética , Hormonas Peptídicas/química , Hormonas Peptídicas/metabolismo , Hormonas Peptídicas/farmacología , Reguladores del Crecimiento de las Plantas/metabolismo , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Proteínas de Plantas/farmacología , Unión Proteica , Proteínas Quinasas/química , Proteínas Quinasas/metabolismo , Multimerización de Proteína/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , Estabilidad Proteica , Estructura Secundaria de Proteína/efectos de los fármacos , Estructura Terciaria de Proteína/efectos de los fármacos , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/metabolismo , Especificidad por SustratoRESUMEN
Plants and animals employ innate immune systems to cope with microbial infection. Pattern-triggered immunity relies on the recognition of microbe-derived patterns by pattern recognition receptors (PRRs). Necrosis and ethylene-inducing peptide 1-like proteins (NLPs) constitute plant immunogenic patterns that are unique, as these proteins are produced by multiple prokaryotic (bacterial) and eukaryotic (fungal, oomycete) species. Here we show that the leucine-rich repeat receptor protein (LRR-RP) RLP23 binds in vivo to a conserved 20-amino-acid fragment found in most NLPs (nlp20), thereby mediating immune activation in Arabidopsis thaliana. RLP23 forms a constitutive, ligand-independent complex with the LRR receptor kinase (LRR-RK) SOBIR1 (Suppressor of Brassinosteroid insensitive 1 (BRI1)-associated kinase (BAK1)-interacting receptor kinase 1), and recruits a second LRR-RK, BAK1, into a tripartite complex upon ligand binding. Stable, ectopic expression of RLP23 in potato (Solanum tuberosum) confers nlp20 pattern recognition and enhanced immunity to destructive oomycete and fungal plant pathogens, such as Phytophthora infestans and Sclerotinia sclerotiorum. PRRs that recognize widespread microbial patterns might be particularly suited for engineering immunity in crop plants.
RESUMEN
The endogenous peptides AtPep1-8 in Arabidopsis mature from the conserved C-terminal portions of their precursor proteins PROPEP1-8, respectively. The two homologous leucine-rich repeat-receptor kinases (LRR-RKs) PEPR1 and PEPR2 act as receptors of AtPeps. AtPep binding leads to stable association of PEPR1,2 with the shared receptor LRR-RK BAK1, eliciting immune responses similar to those induced by pathogens. Here we report a crystal structure of the extracellular LRR domain of PEPR1 (PEPR1LRR) in complex with AtPep1. The structure reveals that AtPep1 adopts a fully extended conformation and binds to the inner surface of the superhelical PEPR1LRR. Biochemical assays showed that AtPep1 is capable of inducing PEPR1LRR-BAK1LRR heterodimerization. The conserved C-terminal portion of AtPep1 dominates AtPep1 binding to PEPR1LRR, with the last amino acid of AtPep1 Asn23 forming extensive interactions with PEPR1LRR. Deletion of the last residue of AtPep1 significantly compromised AtPep1 interaction with PEPR1LRR. Together, our data reveal a conserved structural mechanism of AtPep1 recognition by PEPR1, providing significant insight into prediction of recognition of other peptides by their cognate LRR-RKs.