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Phytophthora sojae is a type of pathogenic oomycete that causes Phytophthora root stem rot (PRSR), which can seriously affect the soybean yield and quality. To subvert immunity, P. sojae secretes a large quantity of effectors. However, the molecular mechanisms regulated by most P. sojae effectors, and their host targets remain unexplored. Previous studies have shown that the expression of PsAvh113, an effector secreted by Phytophthora sojae, enhances viral RNA accumulations and symptoms in Nicotiana benthamiana via VIVE assay. In this study, we analyzed RNA-sequencing data based on disease symptoms in N. benthamiana leaves that were either mocked or infiltrated with PVX carrying the empty vector (EV) and PsAvh113. We identified 1769 differentially expressed genes (DEGs) dependent on PsAvh113. Using stricter criteria screening and Gene Ontology (GO) and Kyoto Encyclopaedia of Genes and Genomes (KEGG) analysis of DEGs, we found that 38 genes were closely enriched in response to PsAvh113 expression. We selected three genes of N. benthamiana (NbNAC86, NbMyb4, and NbERF114) and found their transcriptional levels significantly upregulated in N. benthamiana infected with PVX carrying PsAvh113. Furthermore, individual silencing of these three genes promoted P. capsici infection, while their overexpression increased resistance to P. capsici in N. benthamiana. Our results show that PsAvh113 interacts with transcription factors NbMyb4 and NbERF114 in vivo. Collectively, these data may help us understand the pathogenic mechanism of effectors and manage PRSR in soybeans.
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Antimicrobial acroautophagy/autophagy plays a vital role in degrading intracellular pathogens or microbial molecules in host-microbe interactions. However, microbes evolved various mechanisms to hijack or modulate autophagy to escape elimination. Vector-transmitted phloem-limited bacteria, Candidatus Liberibacter (Ca. Liberibacter) species, cause Huanglongbing (HLB), one of the most catastrophic citrus diseases worldwide, yet contributions of autophagy to HLB disease proliferation remain poorly defined. Here, we report the identification of a virulence effector in "Ca. Liberibacter asiaticus" (Las), SDE3, which is highly conserved among the "Ca. Liberibacter". SDE3 expression not only promotes the disease development of HLB and canker in sweet orange (Citrus sinensis) plants but also facilitates Phytophthora and viral infections in Arabidopsis, and Nicotiana benthamiana (N. benthamiana). SDE3 directly associates with citrus cytosolic glyceraldehyde-3-phosphate dehydrogenases (CsGAPCs), which negatively regulates plant immunity. Overexpression of CsGAPCs and SDE3 significantly inhibits autophagy in citrus, Arabidopsis, and N. benthamiana. Intriguingly, SDE3 undermines autophagy-mediated immunity by the specific degradation of CsATG8 family proteins in a CsGAPC1-dependent manner. CsATG8 degradation is largely rescued by treatment with an inhibitor of the late autophagic pathway, E64d. Furthermore, ectopic expression of CsATG8s enhances Phytophthora resistance. Collectively, these results suggest that SDE3-CsGAPC interactions modulate CsATG8-mediated autophagy to enhance Las progression in citrus.Abbreviations: ACP: asian citrus psyllid; ACD2: ACCELERATED CELL DEATH 2; ATG: autophagy related; Ca. Liberibacter: Candidatus Liberibacter; CaMV: cauliflower mosaic virus; CMV: cucumber mosaic virus; Cs: Citrus sinensis; EV: empty vector; GAPC: cytosolic glyceraldehyde-3-phosphate dehydrogenase; HLB: huanglongbing; H2O2: hydrogen peroxide; Las: liberibacter asiaticus; Laf: liberibacter africanus; Lam: liberibacter americanus; Pst: Pseudomonas syringae pv. tomato; PVX: potato virus X; ROS: reactive oxygen species; SDE3: sec-delivered effector 3; TEM: transmission electron microscopy; VIVE : virus-induced virulence effector; WT: wild-type; Xcc: Xanthomonas citri subsp. citri.
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Arabidopsis , Citrus , Hemípteros , Rhizobiaceae , Animales , Citrus/microbiología , Liberibacter , Peróxido de Hidrógeno , Hemípteros/fisiología , Autofagia , Enfermedades de las Plantas/microbiologíaRESUMEN
Lesion mimic mutants (LMMs) are valuable genetic resources for unraveling plant defense responses including programmed cell death. Here, we identified a rice (Oryza sativa) LMM, spotted leaf 38 (spl38), and demonstrated that spl38 is essential for the formation of hypersensitive response-like lesions and innate immunity. Map-based cloning revealed that SPL38 encodes MEDIATOR SUBUNIT 16 (OsMED16). The spl38 mutant showed enhanced resistance to rice pathogens Magnaporthe oryzae and Xanthomonas oryzae pv. oryzae (Xoo) and exhibited delayed flowering, while OsMED16-overexpressing plants showed increased rice susceptibility to M. oryzae. The OsMED16-edited rice lines were phenotypically similar to the spl38 mutant but were extremely weak, exhibited growth retardation, and eventually died. The C-terminus of OsMED16 showed interaction with the positive immune regulator PATHOGENESIS RELATED 3 (OsPR3), resulting in the competitive repression of its chitinase and chitin-binding activities. Furthermore, the ospr3 osmed16 double mutants did not exhibit the lesion mimic phenotype of the spl38 mutant. Strikingly, OsMED16 exhibited an opposite function in plant defense relative to that of Arabidopsis (Arabidopsis thaliana) AtMED16, most likely because of 2 amino acid substitutions between the monocot and dicot MED16s tested. Collectively, our findings suggest that OsMED16 negatively regulates cell death and immunity in rice, probably via the OsPR3-mediated chitin signaling pathway.
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Oryza , Xanthomonas , Proteínas de Plantas/metabolismo , Inmunidad Innata , Muerte Celular/genética , Apoptosis , Xanthomonas/fisiología , Enfermedades de las Plantas/genética , Regulación de la Expresión Génica de las Plantas , Resistencia a la Enfermedad/genéticaRESUMEN
Phytophthora effector PSR1 suppresses small RNA (sRNA)-mediated immunity in plants, but the underlying mechanism remains unknown. Here, we show that Phytophthora suppressor of RNA silencing 1 (PSR1) contributes to the pathogenicity of Phytophthora sojae and specifically binds to three conserved C-terminal domains of the eukaryotic PSR1-Interacting Protein 1 (PINP1). PINP1 encodes PRP16, a core pre-mRNA splicing factor that unwinds RNA duplexes and binds to primary microRNA transcripts and general RNAs. Intriguingly, PSR1 decreased both RNA helicase and RNA-binding activity of PINP1, thereby dampening sRNA biogenesis and RNA metabolism. The PSR1-PINP1 interaction caused global changes in alternative splicing (AS). A total of 5,135 genes simultaneously exhibited mis-splicing in both PSR1-overexpressing and PINP1-silenced plants. AS upregulated many mRNA transcripts that had their introns retained. The high occurrence of intron retention in AS-induced transcripts significantly promoted Phytophthora pathogen infection in Nicotiana benthamiana, and this might be caused by the production of truncated proteins. Taken together, our findings reveal a key role for PINP1 in regulating sRNA biogenesis and plant immunity.
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Phytophthora , ARN Pequeño no Traducido , Enfermedades de las Plantas , Inmunidad de la Planta , Plantas , Precursores del ARN , Glycine maxRESUMEN
The plant-specific lateral organ boundaries (LOB) domain (LBD) proteins, a family of transcription factors, play important roles in plant growth and development, as well as in responses to various stresses. However, little is known about the functions of LBD genes in soybean (Glycine max). In this study, we investigated the evolution and classification of the LBD family in soybean by a phylogenetic tree of the LBD gene family from 16 species. Phylogenetic analysis categorized these proteins into two classes (Class I and Class II) with seven subgroups. Moreover, we found that all the 18 LBD ancestors in angiosperm were kept in soybean, common bean genomes, and genome-wide duplication, suggesting the main force for the expansion of LBD from common bean to soybean. Analysis of gene expression profiling data indicated that 16 GmLBD genes were significantly induced at different time points after inoculation of soybean plants (cv. Huachun 6) with Phytophthora sojae (P. sojae). We further assessed the role of four highly upregulated genes, GmLBD9, GmLBD16, GmLBD23, and GmLBD88, in plant defense in soybean hairy roots using the transient overexpression and knockdown assays. The results showed that GmLBD9 and GmLBD23 negatively regulate plant immunity against P. sojae, whereas GmLBD16 and GmLBD88 positively manipulate plant immunity against P. sojae. Collectively, our findings expand our knowledge of the origin and evolution of the GmLBD gene family in soybean and promote the potential application of these genes in soybean genetic improvement.
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The soilborne oomycete Phytophthora capsici is the most destructive pathogen of vegetable crops and is responsible for substantial economic losses worldwide. Here, we present an improved genome assembly of P. capsici generated by Oxford Nanopore long-read sequencing (for de novo assembly) and Illumina short-read sequencing (for polishing). The genome of P. capsici is 100.5 Mb in length (GC content = 50.8%) and contains 26,069 predicted protein-coding genes. The whole genome of P. capsici is assembled into 194 scaffolds, 90% of which are larger than 300 kb. The N50 scaffold length and maximum scaffold length are 1.0 and 4.1 Mb, respectively. The whole-genome sequence of P. capsici will broaden our knowledge of this pathogen and enhance our understanding of the molecular basis of its pathogenicity, which will facilitate the development of effective management strategies.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.
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Phytophthora , Genoma , Secuenciación de Nucleótidos de Alto Rendimiento , Phytophthora/genética , VirulenciaRESUMEN
Plant pathogens deliver virulence effectors into plant cells to modulate plant immunity and facilitate infection. Although species-specific virulence effector screening approaches have been developed for several pathogens, these assays do not apply to pathogens that cannot be cultured and/or transformed outside of their hosts. Here, we established a rapid and parallel screening assay, called the virus-induced virulence effector (VIVE) assay, to identify putative effectors in various plant pathogens, including unculturable pathogens, using a virus-based expression vector. The VIVE assay uses the potato virus X (PVX) vector to transiently express candidate effector genes of various bacterial and fungal pathogens into Nicotiana benthamiana leaves. Using the VIVE assay, we successfully identified Avh148 as a potential virulence effector of Phytophthora sojae. Plants infected with PVX carrying Avh148 showed strong viral symptoms and high-level Avh148 and viral RNA accumulation. Analysis of P. sojae Avh148 deletion mutants and soybean hairy roots overexpressing Avh148 revealed that Avh148 is required for full pathogen virulence. In addition, the VIVE assay was optimized in N. benthamiana plants at different developmental stages across a range of Agrobacterium cell densities. Overall, we identified six novel virulence effectors from seven pathogens, thus demonstrating the broad effectiveness of the VIVE assay in plant pathology research.
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Glycine max/virología , Nicotiana/virología , Phytophthora/genética , Enfermedades de las Plantas/virología , Potexvirus/genética , Factores de Virulencia/genética , Phytophthora/patogenicidad , Enfermedades de las Plantas/inmunología , Enfermedades de las Plantas/parasitología , Inmunidad de la Planta , Hojas de la Planta/genética , Hojas de la Planta/inmunología , Hojas de la Planta/parasitología , Hojas de la Planta/virología , Raíces de Plantas/genética , Raíces de Plantas/inmunología , Raíces de Plantas/parasitología , Raíces de Plantas/virología , ARN Viral/genética , Eliminación de Secuencia , Glycine max/genética , Glycine max/inmunología , Glycine max/parasitología , Nicotiana/genética , Nicotiana/inmunología , Nicotiana/parasitología , VirulenciaRESUMEN
Over the past 30 years, human disturbance and habitat fragmentation have severely endangered the survival of common wild rice (Oryza rufipogon Griff.) in China. A better understanding of the genetic structure of O. rufipogon populations will therefore be useful for the development of conservation strategies. We examined the diversity and genetic structure of natural O. rufipogon populations at the national, provincial, and local levels using simple sequence repeat (SSR) markers. Twenty representative populations from sites across China showed high levels of genetic variability, and approximately 44% of the total genetic variation was among populations. At the local level, we studied fourteen populations in Guangxi Province and four populations in Jiangxi Province. Populations from similar ecosystems showed less genetic differentiation, and local environmental conditions rather than geographic distance appeared to have influenced gene flow during population genetic evolution. We identified a triangular area, including northern Hainan, southern Guangdong, and southwestern Guangxi, as the genetic diversity center of O. rufipogon in China, and we proposed that this area should be given priority during the development of ex situ and in situ conservation strategies. Populations from less common ecosystem types should also be given priority for in situ conservation.
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Productos Agrícolas/genética , Especies en Peligro de Extinción , Genes de Plantas , Variación Genética , Oryza/genética , Alelos , China , ADN de Plantas/genética , Ecosistema , Evolución Molecular , Flujo Génico , Repeticiones de Microsatélite/genética , Reacción en Cadena de la Polimerasa , Análisis de Componente PrincipalRESUMEN
RNA silencing is an evolutionarily conserved, sequence-specific gene regulation mechanism in eukaryotes. Several plant pathogens have evolved proteins with the ability to inhibit the host plant RNA silencing pathway. Unlike virus effector proteins, only several secreted effector proteins have showed the ability to suppress RNA silencing in bacterial, oomycete, and fungal pathogens, and the molecular functions of most effectors remain largely unknown. Here, we describe in detail a slightly modified version of the co-infiltration assay that could serve as a general method for observing RNA silencing and for characterizing effector proteins secreted by plant pathogens. The key steps of the approach are choosing the healthy and fully developed leaves, adjusting the bacteria culture to the appropriate optical density (OD) at 600 nm, and observing green fluorescent protein (GFP) fluorescence at the optimum time on the infiltrated leaves in order to avoid omitting effectors with weak suppression activity. This improved protocol will contribute to rapid, accurate, and extensive screening of RNA silencing suppressors and serve as an excellent starting point for investigating the molecular functions of these proteins.
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Enfermedades de las Plantas/genética , Plantas/genética , Interferencia de ARN , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Hojas de la Planta/metabolismo , Plantas/metabolismoRESUMEN
Phytophthora pathogens manipulate host innate immunity by secreting numerous RxLR effectors, thereby facilitating pathogen colonization. Predicted single and tandem repeats of WY domains are the most prominent C-terminal motifs conserved across the Phytophthora RxLR superfamily. However, the functions of individual WY domains in effectors remain poorly understood. The Phytophthora sojae effector PSR1 promotes infection by suppressing small RNA biogenesis in plant hosts. We identified one single WY domain following the RxLR motif in PSR1. This domain was required for RNA silencing suppression activity and infection in Nicotiana benthamiana, Arabidopsis and soybean. Mutations of the conserved residues in the WY domain did not affect the subcellular localization of PSR1 but abolished its effect on plant development and resistance to viral and Phytophthora pathogens. This is at least in part due to decreased protein stability of the PSR1 mutants in planta. The identification of the WY domain in PSR1 allows predicts that a family of PSR1-like effectors also possess RNA silencing suppression activity. Mutation of the conserved residues in two members of this family, PpPSR1L from P. parasitica and PcPSR1L from P. capsici, perturbed their biological functions, indicating that the WY domain is critical in Phytophthora PSR1 and PSR1-like effectors.
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Phytophthora/metabolismo , Proteínas/química , Proteínas/metabolismo , Interferencia de ARN , Secuencia de Aminoácidos , Arabidopsis/crecimiento & desarrollo , Arabidopsis/microbiología , Secuencia Conservada , Mutación/genética , Fenotipo , Phytophthora/patogenicidad , Raíces de Plantas/microbiología , Unión Proteica , Dominios Proteicos , Proteínas/genética , Glycine max/microbiologíaRESUMEN
The citrus industry is facing an unprecedented challenge from Huanglongbing (HLB). All cultivars can be affected by the HLB-associated bacterium 'Candidatus Liberibacter asiaticus' (CLas) and there is no known resistance. Insight into HLB pathogenesis is urgently needed in order to develop effective management strategies. Here, we use Sec-delivered effector 1 (SDE1), which is conserved in all CLas isolates, as a molecular probe to understand CLas virulence. We show that SDE1 directly interacts with citrus papain-like cysteine proteases (PLCPs) and inhibits protease activity. PLCPs are defense-inducible and exhibit increased protein accumulation in CLas-infected trees, suggesting a role in citrus defense responses. We analyzed PLCP activity in field samples, revealing specific members that increase in abundance but remain unchanged in activity during infection. SDE1-expressing transgenic citrus also exhibit reduced PLCP activity. These data demonstrate that SDE1 inhibits citrus PLCPs, which are immune-related proteases that enhance defense responses in plants.
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Citrus/microbiología , Proteasas de Cisteína/genética , Inhibidores de Cisteína Proteinasa/metabolismo , Evasión Inmune , Enfermedades de las Plantas/microbiología , Proteínas de Plantas/antagonistas & inhibidores , Rhizobiaceae/patogenicidad , Proteínas Bacterianas/biosíntesis , Proteínas Bacterianas/genética , Citrus/clasificación , Citrus/genética , Citrus/inmunología , Proteasas de Cisteína/inmunología , Inhibidores de Cisteína Proteinasa/química , Regulación de la Expresión Génica , Filogenia , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/inmunología , Inmunidad de la Planta/genética , Hojas de la Planta/genética , Hojas de la Planta/inmunología , Hojas de la Planta/microbiología , Proteínas de Plantas/genética , Proteínas de Plantas/inmunología , Rhizobiaceae/genética , Factores de Virulencia/biosíntesis , Factores de Virulencia/genéticaRESUMEN
The citrus industry is facing an unprecedented crisis due to Huanglongbing (HLB, aka citrus greening disease), a bacterial disease associated with the pathogen Candidatus Liberibacter asiaticus (CLas) that affects all commercial varieties. Transmitted by the Asian citrus psyllid (ACP), CLas colonizes citrus phloem, leading to reduced yield and fruit quality, and eventually tree decline and death. Since adequate curative measures are not available, a key step in HLB management is to restrict the spread of the disease by identifying infected trees and removing them in a timely manner. However, uneven distribution of CLas cells in infected trees and the long latency for disease symptom development makes sampling of trees for CLas detection challenging. Here, we report that a CLas secreted protein can be used as a biomarker for detecting HLB infected citrus. Proteins secreted from CLas cells can presumably move along the phloem, beyond the site of ACP inoculation and CLas colonized plant cells, thereby increasing the chance of detecting infected trees. We generated a polyclonal antibody that effectively binds to the secreted protein and developed serological assays that can successfully detect CLas infection. This work demonstrates that antibody-based diagnosis using a CLas secreted protein as the detection marker for infected trees offers a high-throughput and economic approach that complements the approved quantitative polymerase chain reaction-based methods to enhance HLB management programs.
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A broad range of parasites rely on the functions of effector proteins to subvert host immune response and facilitate disease development. The notorious Phytophthora pathogens evolved effectors with RNA silencing suppression activity to promote infection in plant hosts. Here we report that the Phytophthora Suppressor of RNA Silencing 1 (PSR1) can bind to an evolutionarily conserved nuclear protein containing the aspartate-glutamate-alanine-histidine-box RNA helicase domain in plants. This protein, designated PSR1-Interacting Protein 1 (PINP1), regulates the accumulation of both microRNAs and endogenous small interfering RNAs in Arabidopsis. A null mutation of PINP1 causes embryonic lethality, and silencing of PINP1 leads to developmental defects and hypersusceptibility to Phytophthora infection. These phenotypes are reminiscent of transgenic plants expressing PSR1, supporting PINP1 as a direct virulence target of PSR1. We further demonstrate that the localization of the Dicer-like 1 protein complex is impaired in the nucleus of PINP1-silenced or PSR1-expressing cells, indicating that PINP1 may facilitate small RNA processing by affecting the assembly of dicing complexes. A similar function of PINP1 homologous genes in development and immunity was also observed in Nicotiana benthamiana. These findings highlight PINP1 as a previously unidentified component of RNA silencing that regulates distinct classes of small RNAs in plants. Importantly, Phytophthora has evolved effectors to target PINP1 in order to promote infection.
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Arabidopsis/genética , Arabidopsis/parasitología , Phytophthora/metabolismo , ARN Interferente Pequeño/genética , Alelos , Núcleo Celular/metabolismo , Silenciador del Gen , MicroARNs/metabolismo , Fenotipo , Enfermedades de las Plantas/parasitología , Plantas Modificadas Genéticamente , Estructura Terciaria de Proteína , ARN Helicasas/metabolismo , Interferencia de ARN , ARN de Planta/genética , Nicotiana/metabolismo , VirulenciaRESUMEN
Citrus stubborn disease (CSD), first identified in California, is a widespread bacterial disease found in most arid citrus-producing regions in the United States and the Mediterranean Region. The disease is caused by Spiroplasma citri, an insect-transmitted and phloem-colonizing bacterium. CSD causes significant tree damage resulting in loss of fruit production and quality. Detection of CSD is challenging due to low and fluctuating titer and sporadic distribution of the pathogen in infected trees. In this study, we report the development of a novel diagnostic method for CSD using an S. citri-secreted protein as the detection marker. Microbial pathogens secrete a variety of proteins during infection that can potentially disperse systemically in infected plants with the vascular flow. Therefore, their distribution may not be restricted to the pathogen infection sites and could be used as a biological marker for infection. Using mass spectrometry analysis, we identified a unique secreted protein from S. citri that is highly expressed in the presence of citrus phloem extract. ScCCPP1, an antibody generated against this protein, was able to distinguish S. citri-infected citrus and periwinkle from healthy plants. In addition, the antiserum could be used to detect CSD using a simple direct tissue print assay without the need for sample processing or specialized lab equipment and may be suitable for field surveys. This study provides proof of a novel concept of using pathogen-secreted protein as a marker for diagnosis of a citrus bacterial disease and can probably be applied to other plant diseases.
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Anticuerpos Antibacterianos/inmunología , Especificidad de Anticuerpos , Proteínas Bacterianas/aislamiento & purificación , Citrus/microbiología , Enfermedades de las Plantas/microbiología , Spiroplasma/inmunología , Proteínas Bacterianas/inmunología , Proteínas Bacterianas/metabolismo , Biomarcadores/metabolismo , Citrus/química , Espectrometría de Masas , Floema/química , Floema/microbiología , Extractos Vegetales/farmacología , Hojas de la Planta/química , Hojas de la Planta/microbiología , Tallos de la Planta/química , Tallos de la Planta/microbiología , Especificidad de la Especie , Spiroplasma/efectos de los fármacos , Spiroplasma/aislamiento & purificación , Vinca/microbiologíaRESUMEN
OBJECTIVE: This study was to screen for the miRNAs differently expressed in peripheral blood mononuclear cells (PBMC) of RA, to further identify the expression of miR-155 in RA PBMC and fibroblast-like synoviocytes (FLS), and to evaluate the function of miR-155 in RA-FLS. METHODS: Microarray was used to screen for differentially expressed miRNAs in RA PBMC. miR-155 expression in PBMC and FLS of RA were identified by real-time PCR. Enforced overexpression and downexpression of miR-155 were used to investigate the function of miR-155 in RA-FLS. Expression of IKBKE which was previously identified as the actual target of miR-155 was examined by Western blot and real-time PCR in RA-FLS. RESULTS: miR-155 levels were increased in both PBMC and FLS of RA and could be induced by TNF- α . Upregulation of miR-155 decreased MMP-3 levels and suppressed proliferation and invasion of RA-FLS. Inverse relationship between the expressions of miR-155 and the MMPs production-related protein IKBKE was found. CONCLUSION: An inflammatory milieu may alter miRNA expression profiles in rheumatoid arthritis. miR-155 is upregulated in RA-FLS, and it may be a protective factor against the inflammatory effect in part by attenuating expression of IKBKE.
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Artritis Reumatoide/genética , Fibroblastos/metabolismo , Regulación de la Expresión Génica , Leucocitos Mononucleares/metabolismo , MicroARNs/genética , Membrana Sinovial/citología , Adulto , Apoptosis/genética , Estudios de Casos y Controles , Proliferación Celular , Femenino , Fibroblastos/efectos de los fármacos , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Quinasa I-kappa B/genética , Quinasa I-kappa B/metabolismo , Leucocitos Mononucleares/efectos de los fármacos , Masculino , Metaloproteinasa 3 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , MicroARNs/metabolismo , Persona de Mediana Edad , Reproducibilidad de los Resultados , Factor de Crecimiento Transformador beta/biosíntesis , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Regulación hacia Arriba , Adulto JovenRESUMEN
Effectors are essential virulence proteins produced by a broad range of parasites, including viruses, bacteria, fungi, oomycetes, protozoa, insects and nematodes. Upon entry into host cells, pathogen effectors manipulate specific physiological processes or signaling pathways to subvert host immunity. Most effectors, especially those of eukaryotic pathogens, remain functionally uncharacterized. Here, we show that two effectors from the oomycete plant pathogen Phytophthora sojae suppress RNA silencing in plants by inhibiting the biogenesis of small RNAs. Ectopic expression of these Phytophthora suppressors of RNA silencing enhances plant susceptibility to both a virus and Phytophthora, showing that some eukaryotic pathogens have evolved virulence proteins that target host RNA silencing processes to promote infection. These findings identify RNA silencing suppression as a common strategy used by pathogens across kingdoms to cause disease and are consistent with RNA silencing having key roles in host defense.
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Phytophthora , Enfermedades de las Plantas , Plantas , Interferencia de ARN , Regulación de la Expresión Génica , Interacciones Huésped-Patógeno/genética , Interacciones Huésped-Patógeno/inmunología , Infecciones/genética , Infecciones/parasitología , Phytophthora/genética , Phytophthora/metabolismo , Phytophthora/patogenicidad , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/parasitología , Plantas/genética , Plantas/parasitología , Proteínas/genética , Transducción de Señal , VirulenciaRESUMEN
Architecture of the rice inflorescence, which is determined mainly by the morphology, number and length of primary and secondary inflorescence branches, is an important agronomical trait. In the current study, we characterized a novel dense and erect panicle (EP) mutant, dep3, derived from the Oryza sativa ssp. japonica cultivar Hwacheong treated with N-methyl-N-nitrosourea. The panicle of the dep3 mutant remained erect from flowering to full maturation, whereas the panicle of the wild type plant began to droop after flowering. The dep3 mutation also regulated other panicle characteristics, including panicle length, grain shape and grain number per panicle. Anatomical observations revealed that the dep3 mutant had more small vascular bundles and a thicker culm than wild type plants, explaining the EP phenotype. Genetic analysis indicated that the phenotype with the dense and EP was controlled by a single recessive gene, termed dep3. The DEP3 gene was identified as the candidate via a map-based cloning approach and was predicted to encode a patatin-like phospholipase A2 (PLA2) superfamily domain-containing protein. The mutant allele gene carried a 408 bp genomic deletion within LOC_Os06g46350, which included the last 47 bp coding region of the third exon and the first 361 bp of the 3'-untranslated region. Taken together, our results indicated that the patatin-like PLA2 might play a significant role in the formation of vascular bundles, and that the dep3 mutant may provide another EP resource for rice breeding programs.
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Mapeo Cromosómico , Genes de Plantas , Oryza/genética , Proteínas de Plantas/genética , Semillas/crecimiento & desarrollo , Alelos , Secuencia de Bases , Cruzamiento , Cromosomas de las Plantas , Clonación Molecular , Regulación de la Expresión Génica de las Plantas , Genotipo , Inflorescencia , Datos de Secuencia Molecular , Mutación , Oryza/crecimiento & desarrollo , Reacción en Cadena de la Polimerasa , Semillas/genéticaRESUMEN
Tumor necrosis factor alpha (TNF-alpha) is a major inflammatory mediator that exhibits actions leading to tissue destruction and hampering recovery from damage. At present, two antibodies against human TNF-alpha (hTNF-alpha) are available, which are widely used for the clinic treatment of certain inflammatory diseases. This work was undertaken to identify a novel functional epitope of hTNF-alpha. We performed screening peptide library against anti-hTNF-alpha antibodies, ELISA and competitive ELISA to obtain the epitope of hTNF-alpha. The key residues of the epitope were identified by means of combinatorial alanine scanning and site-specific mutagenesis. The N terminus (80-91 aa) of hTNF-alpha proved to be a novel epitope (YG1). The two amino acids of YG1, proline and valine, were identified as the key residues, which were important for hTNF-alpha biological function. Furthermore, the function of the epitope was addressed on an animal model of collagen-induced arthritis (CIA). CIA could be suppressed in an animal model by prevaccination with the derivative peptides of YG1. The antibodies of YG1 could also inhibit the cytotoxicity of hTNF-alpha. These results demonstrate that YG1 is a novel epitope associated with the biological function of hTNF-alpha and the antibodies against YG1 can inhibit the development of CIA in animal model, so it would be a potential target of new therapeutic antibodies.
Asunto(s)
Anticuerpos/inmunología , Artritis Experimental/prevención & control , Colágeno/administración & dosificación , Epítopos/inmunología , Factor de Necrosis Tumoral alfa/inmunología , Secuencia de Aminoácidos , Animales , Artritis Experimental/inducido químicamente , Secuencia de Bases , Cartilla de ADN , Ensayo de Inmunoadsorción Enzimática , Epítopos/química , Humanos , Ratones , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Conejos , Homología de Secuencia de Aminoácido , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
Rheumatoid arthritis (RA) is a systemic autoimmune disease mediated by T cells. Collagen type II (CII) is one of the autoantigens associated with RA. CII263-272 is a predominant CII antigenic peptide that can induce T-cell activation upon binding to MHC and interaction with the appropriate T-cell receptor (TCR). Altered CII263-272 peptides with substitution of specific amino acids could bind to RA-associated HLA-DR4/1 with no T cell stimulating effects and could inhibit T cell activation in RA. We performed this study to evaluate the effect of mucosal administration and to explore the mechanism of the inhibitory effect of altered CII263-272 peptide (267Q-->A, 270K-->A and 271G-->A) on collagen induced arthritis (CIA). CIA was induced in Lewis rats by immunization with bovine CII. Altered CII263-272 peptide was given intranasally beginning from arthritis onset. Wild CII263-272 peptide or PBS was administered as controls. Therapeutic effects were evaluated by arthritis scores, body weight change, and joint pathologic scores. The anti-CII antibody and its subtypes and the cytokines, IFN-gamma, IL-10, and IL-17 were measured with ELISA. Foxp3+CD4+CD25+ regulatory T cell induction was assessed by FACS analysis. Following treatment with the altered CII263-272 peptide, arthiritis scores were reduced and body weight was increased. The altered CII263-272 peptide could retard the histologic lesion of the joints. The titers of anti-CII antibodies IgG2a in altered CII263-272 peptide treated rats decreased markedly compared to PBS-treated rats. The serum levels of IFN-gamma in rats treated with altered peptide was lower than that of rats treated with wild CII263-272 peptide and PBS. No differences were observed in the levels of serum IL-10 among the three groups. The altered CII263-272 peptide could decrease serum level of IL-17 and increase peripheral Foxp3+CD4+CD25+ T cells at early stage of CIA. Mucosal administration of altered CII263-272 peptide could effectively inhibit the progression of CIA. Altered CII263-272 peptide could suppress Th17 cells and expand regulatory T cells in the early stage of the disease. The IgG2a subtype of anti-CII antibodies and IFN-gamma were reduced and in vivo Th1 responses were inhibited as a result of altered CII peptide treatment. Altered CII peptide is likely therapeutic in RA.
Asunto(s)
Artritis Experimental/inmunología , Artritis Experimental/prevención & control , Activación de Linfocitos/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Linfocitos T Reguladores/efectos de los fármacos , Células TH1/efectos de los fármacos , Administración Intranasal , Animales , Artritis Experimental/patología , Citocinas/metabolismo , Femenino , Miembro Posterior , Articulaciones/efectos de los fármacos , Articulaciones/patología , Articulaciones/fisiopatología , Mucosa Nasal/efectos de los fármacos , Fragmentos de Péptidos/administración & dosificación , Fragmentos de Péptidos/inmunología , Ratas , Ratas Endogámicas Lew , Índice de Severidad de la Enfermedad , Organismos Libres de Patógenos Específicos , Linfocitos T Reguladores/inmunología , Células TH1/inmunologíaRESUMEN
To investigate the specificity, sensitivity, and concomitant presence of antibodies against histones (H), histone H1 (H1), and histone H3 (H3) in patients with systemic lupus erythematosus (SLE) and analyze their association with SLE. Serum IgG anti-histones antibodies were detected by enzyme-linked immunosorbent assay in 144 SLE patients consisting of 24 neuropsychiatric lupus (NPSLE), 65 lupus nephritis (LN), and 55 SLE, 100 other rheumatic diseases patients, as well as 40 healthy controls. Clinical and biological parameters of the patients were also evaluated. Anti-H, anti-H1, and anti-H3 antibodies yielded a sensitivity of approximately 33% and a specificity of more than 93% for SLE, which was comparable to that found for anti-double-stranded DNA (anti-dsDNa) antibodies. More significantly, anti-histone antibody is found in approximately 50% of patients with NPSLE compared with LN. Moreover, the titers of anti-histones antibodies of NPSLE patients were significantly higher than that of patients with SLE and LN. The sequential analysis revealed a close correlation of anti-H and anti-H1 antibodies with SLE disease activity. There was an approximate 30% positive rate of anti-histones antibodies in 144 SLE patients lacking anti-nucleosome, anti-mDNA, anti-Sm, and anti-dsDNA antibodies. Antibodies to histones H1 and H3 are markers with high specificity of 93.6-96.4% for SLE. The anti-histone antibody markers are prevalent in approximately 50% of NPSLE. Furthermore, there was a strong correlation with SLE disease activity index and levels of antibodies to histones and H1.