RESUMEN
As a core subunit of the SCF complex that promotes protein degradation through the 26S proteasome, S-phase kinase-associated protein 1 (SKP1) plays important roles in multiple cellular processes in eukaryotes, including gibberellin (GA), jasmonate, ethylene, auxin and light responses. P7-2 encoded by Rice black streaked dwarf virus (RBSDV), a devastating viral pathogen that causes severe symptoms in infected plants, interacts with SKP1 from different plants. However, whether RBSDV P7-2 forms a SCF complex and targets host proteins is poorly understood. In this study, we conducted yeast two-hybrid assays to further explore the interactions between P7-2 and 25 type I Oryza sativa SKP1-like (OSK) proteins, and found that P7-2 interacted with eight OSK members with different binding affinity. Co-immunoprecipitation assay further confirmed the interaction of P7-2 with OSK1, OSK5 and OSK20. It was also shown that P7-2, together with OSK1 and O. sativa Cullin-1, was able to form the SCF complex. Moreover, yeast two-hybrid assays revealed that P7-2 interacted with gibberellin insensitive dwarf2 (GID2) from rice and maize plants, which is essential for regulating the GA signaling pathway. It was further demonstrated that the N-terminal region of P7-2 was necessary for the interaction with GID2. Overall, these results indicated that P7-2 functioned as a component of the SCF complex in rice, and interaction of P7-2 with GID2 implied possible roles of the GA signaling pathway during RBSDV infection.
Asunto(s)
Giberelinas/metabolismo , Complejos Multiproteicos/metabolismo , Oryza/metabolismo , Oryza/virología , Proteínas de Plantas/metabolismo , Reoviridae/metabolismo , Proteínas Quinasas Asociadas a Fase-S/metabolismo , Proteínas Virales/metabolismo , Inmunoprecipitación , Hojas de la Planta/metabolismo , Unión Proteica , Reproducibilidad de los Resultados , Nicotiana/metabolismo , Técnicas del Sistema de Dos Híbridos , Zea maysRESUMEN
Gibberellin (GA) 20-oxidase (GA20ox) catalyses consecutive steps of oxidation in the late part of the GA biosynthetic pathway. A T-DNA insertion mutant (17S-14) in rice, with an elongated phenotype, was isolated. Analysis of the flanking sequences of the T-DNA insertion site revealed that an incomplete T-DNA integration resulted in enhanced constitutively expression of downstream OsGA20ox3 in the mutant. The accumulation of bioactive GA(1) and GA(4) were increased in the mutant in comparison with the wild-type plant. Transgenic plants overexpressing OsGA20ox3 showed phenotypes similar to those of the 17S-14 mutant, and the RNA interference (RNAi) lines that had decreased OsGA20ox3 expression exhibited a semidwarf phenotype. Expression of OsGA20ox3 was detected in the leaves and roots of young seedlings, immature panicles, anthers, and pollens, based on ß-glucuronidase (GUS) activity staining in transgenic plants expressing the OsGA20ox3 promoter fused to the GUS gene. The OsGA20ox3 RNAi lines showed enhanced resistance against rice pathogens Magnaporthe oryzae (causing rice blast) and Xanthomonas oryzae pv. oryzae (causing bacterial blight) and increased expression of defense-related genes. Conversely, OsGA20ox3-overexpressing plants were more susceptible to these pathogens comparing with the wild-type plants. The susceptibility of wild-type plants to X. oryzae pv. oryzae was increased by exogenous application of GA(3) and decreased by S-3307 treatment. Together, the results provide direct evidence for a critical role of OsGA20ox3 in regulating not only plant stature but also disease resistance in rice.
Asunto(s)
Giberelinas/metabolismo , Magnaporthe/patogenicidad , Oxigenasas de Función Mixta/metabolismo , Oryza/enzimología , Enfermedades de las Plantas/inmunología , Xanthomonas/patogenicidad , ADN Bacteriano , Resistencia a la Enfermedad , Flores , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Giberelinas/análisis , Oxigenasas de Función Mixta/genética , Mutagénesis Insercional , Oryza/genética , Oryza/inmunología , Oryza/fisiología , Fenotipo , Enfermedades de las Plantas/microbiología , Hojas de la Planta/genética , Hojas de la Planta/inmunología , Hojas de la Planta/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raíces de Plantas , Plantas Modificadas Genéticamente/genética , Plantas Modificadas Genéticamente/inmunología , Plantas Modificadas Genéticamente/metabolismo , Interferencia de ARN , Ácido Salicílico/metabolismo , Plantones/genética , Plantones/inmunología , Plantones/metabolismoRESUMEN
WRKY transcription factors, originally isolated from plants contain one or two conserved WRKY domains, about 60 amino acid residues with the WRKYGQK sequence followed by a C2H2 or C2HC zinc finger motif. Evidence is accumulating to suggest that the WRKY proteins play significant roles in responses to biotic and abiotic stresses, and in development. In this research, we identified 102 putative WRKY genes from the rice genome and compared them with those from Arabidopsis. The WRKY genes from rice and Arabidopsis were divided into three groups with several subgroups on the basis of phylogenies and the basic structure of the WRKY domains (WDs). The phylogenetic trees generated from the WDs and the genes indicate that the WRKY gene family arose during evolution through duplication and that the dramatic amplification of rice WRKY genes in group III is due to tandem and segmental gene duplication compared with those of Arabidopsis. The result suggests that some of the rice WRKY genes in group III are evolutionarily more active than those in Arabidopsis, and may have specific roles in monocotyledonous plants. Further, it was possible to identify the presence of WRKY-like genes in protists (Giardia lamblia and Dictyostelium discoideum) and green algae Chlamydomonas reinhardtii through database research, demonstrating the ancient origin of the gene family. The results obtained by alignments of the WDs from different species and other analysis imply that domain gain and loss is a divergent force for expansion of the WRKY gene family, and that a rapid amplification of the WRKY genes predate the divergence of monocots and dicots. On the basis of these results, we believe that genes encoding a single WD may have been derived from the C-terminal WD of the genes harboring two WDs. The conserved intron splicing positions in the WDs of higher plants offer clues about WRKY gene evolution, annotation, and classification.
Asunto(s)
Arabidopsis/genética , Oryza/genética , Proteínas de Plantas/genética , Factores de Transcripción/genética , Secuencias de Aminoácidos/genética , Secuencia de Aminoácidos , Proteínas de Arabidopsis/genética , Secuencia Conservada , Genoma , Datos de Secuencia Molecular , Familia de Multigenes/genética , Filogenia , Estructura Terciaria de Proteína/genéticaRESUMEN
Osmotin promoter binding protein 1 (OPBP1), an AP2/EREBP-like transcription factor of tobacco (Nicotiana tabacum), was isolated using a yeast one-hybrid system. RNA gel blot analysis indicated that expression of the OPBP1 gene was induced by elicitor cryptogein, NaCl, ethephon, methyl jasmonate, as well as cycloheximide. Transient expression analysis using an OPBP1-eGFP fusion gene in onion epidermal cells revealed that the OPBP1 protein was targeted to the nuclear. Further, electrophoretic mobility shift assays demonstrated that the recombinant OPBP1 protein could bind to an oligonucleotide containing the GCC-box cis element. Transgenic tobacco plants with an over expression of the OPBP1 gene accumulated high levels of PR-1a and PR-5d genes and exhibited enhanced resistance to infection by Pseudomonas syringae pv tabaci and Phytophthora parasitica var nicotianae pathogens. They also exhibited increased tolerance to salt stress. These results suggest that OPBP1 might be a transcriptional regulator capable of regulating expression in sets of stress-related genes.
Asunto(s)
Regulación de la Expresión Génica de las Plantas , Nicotiana/genética , Cloruro de Sodio/farmacología , Factores de Transcripción/genética , Acetatos/farmacología , Adaptación Fisiológica/efectos de los fármacos , Adaptación Fisiológica/genética , Proteínas Algáceas/farmacología , Secuencia de Bases , Sitios de Unión/genética , Unión Competitiva , Núcleo Celular/metabolismo , Cicloheximida/farmacología , Ciclopentanos/farmacología , Proteínas Fúngicas , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Inmunidad Innata/genética , Mutación , Oligonucleótidos/genética , Oligonucleótidos/metabolismo , Compuestos Organofosforados/farmacología , Oxilipinas , Filogenia , Phytophthora/crecimiento & desarrollo , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/microbiología , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Unión Proteica , Pseudomonas syringae/crecimiento & desarrollo , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Nicotiana/efectos de los fármacos , Nicotiana/microbiología , Factores de Transcripción/metabolismoRESUMEN
Lagenaria leucantha is an important vegetable crop and a potential model for the study of fruit development. To study the function of D cyclins in fruit development, full-length cDNA clones for two D cyclin genes were isolated from young ovaries of Lagenaria leucantha. They were classified as D3 cyclins by sequence similarities and phylogenetic analysis, and nominated LlCycD3;1 and LlCycD3;2, respectively. The deduced amino acid sequence of both LlCycD3 genes contained a retinoblastoma-binding motif and a PEST-destruction motif. Unpollinated ovaries failed to develop and eventually aborted. N-(2-chloro-4-pyridyl)-N'-phenylurea (CPPU) induced parthenocarpic fruit significantly larger than pollinated ones. In unpollinated ovaries, the expression of both LlCycD3 genes was abundant at anthesis and then suddenly decreased, concomitant with the cessation of cell division. Pollination/fertilization induced an activation of the cell cycle accompanied by a large increase in the transcript levels of LlCycD3;1 and LlCycD3;2 in young fruits. Treating ovaries with CPPU also reactivated cell division and transcription of CycD3 genes and the effect was more rapid and pronounced than after pollination/fertilization.