RESUMO
Plant pathogens of the oomycete genus Phytophthora produce virulence factors, known as RxLR effector proteins that are transferred into host cells to suppress disease resistance. Here, we analyse the function of the highly conserved RxLR24 effector of Phytophthora brassicae. RxLR24 was expressed early in the interaction with Arabidopsis plants and ectopic expression in the host enhanced leaf colonization and zoosporangia formation. Co-immunoprecipitation (Co-IP) experiments followed by mass spectrometry identified different members of the RABA GTPase family as putative RxLR24 targets. Physical interaction of RxLR24 or its homologue from the potato pathogen Phytophthora infestans with different RABA GTPases of Arabidopsis or potato, respectively, was confirmed by reciprocal Co-IP. In line with the function of RABA GTPases in vesicular secretion, RxLR24 co-localized with RABA1a to vesicles and the plasma membrane. The effect of RxLR24 on the secretory process was analysed with fusion constructs of secreted antimicrobial proteins with a pH-sensitive GFP tag. PATHOGENESIS RELATED PROTEIN 1 (PR-1) and DEFENSIN (PDF1.2) were efficiently exported in control tissue, whereas in the presence of RxLR24 they both accumulated in the endoplasmic reticulum. Together our results imply a virulence function of RxLR24 effectors as inhibitors of RABA GTPase-mediated vesicular secretion of antimicrobial PR-1, PDF1.2 and possibly other defence-related compounds.
Assuntos
Peptídeos Catiônicos Antimicrobianos/metabolismo , Arabidopsis/microbiologia , Vesículas Extracelulares/metabolismo , Proteínas Fúngicas/fisiologia , Phytophthora/fisiologia , Doenças das Plantas/microbiologia , Solanum tuberosum/microbiologia , Fatores de Virulência/fisiologia , Proteínas rab de Ligação ao GTP/metabolismo , Arabidopsis/imunologia , Arabidopsis/fisiologia , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Interações Hospedeiro-Patógeno , Imunoprecipitação , Phytophthora/genética , Phytophthora/metabolismo , Doenças das Plantas/imunologia , Folhas de Planta/microbiologia , Solanum tuberosum/imunologia , Solanum tuberosum/fisiologia , Fatores de Virulência/genética , Fatores de Virulência/metabolismoRESUMO
Pathogen and Circadian Controlled 1 (PCC1) was previously characterized as a regulator of defence against pathogens and stress-activated transition to flowering. Plants expressing an RNA interference construct for the PCC1 gene (iPCC1 plants) showed a pleiotropic phenotype. They were hypersensitive to abscisic acid (ABA) as shown by reduced germination potential and seedling establishment, as well as reduced stomatal aperture and main root length in ABA-supplemented media. In addition, iPCC1 plants displayed alterations in polar lipid contents and their corresponding fatty acids. Importantly, a significant reduction in the content of phosphatidylinositol (PI) was observed in iPCC1 leaves when compared with wild-type plants. A trend in reduced levels of 18:0 and increased levels of 18:2 and particularly 18:3 was also detected in several classes of polar lipids. The enhanced ABA-mediated responses and the reduced content of PI might be responsible for iPCC1 plants displaying a complex pattern of defence against pathogens of different lifestyles. iPCC1 plants were more susceptible to the hemi-biotrophic oomycete pathogen Phytophthora brassicae and more resistant to the necrotrophic fungal pathogen Botrytis cinerea compared with wild-type plants.
Assuntos
Ácido Abscísico/farmacologia , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Arabidopsis/efeitos dos fármacos , Proteínas de Arabidopsis/genética , Resistência à Doença/efeitos dos fármacos , Resistência à Doença/genética , Regulação da Expressão Gênica de Plantas , Fosfatidilinositóis/metabolismo , Doenças das PlantasRESUMO
Plant immune responses to pathogens are often associated with enhanced production of reactive oxygen species (ROS), known as the oxidative burst, and with rapid hypersensitive host cell death (the hypersensitive response, HR) at sites of attempted infection. It is generally accepted that the oxidative burst acts as a promotive signal for HR, and that HR is highly correlated with efficient disease resistance. We have identified the Arabidopsis mutant rph1 (resistance to Phytophthora 1), which is susceptible to the oomycete pathogen Phytophthora brassicae despite rapid induction of HR. The susceptibility of rph1 was specific for P. brassicae and coincided with a reduced oxidative burst, a runaway cell-death response, and failure to properly activate the expression of defence-related genes. From these results, we conclude that, in the immune response to P. brassicae, (i) HR is not sufficient to stop the pathogen, (ii) HR initiation can occur in the absence of a major oxidative burst, (iii) the oxidative burst plays a role in limiting the spread of cell death, and (iv) RPH1 is a positive regulator of the P. brassicae-induced oxidative burst and enhanced expression of defence-related genes. Surprisingly, RPH1 encodes an evolutionary highly conserved chloroplast protein, indicating a function of this organelle in activation of a subset of immune reactions in response to P. brassicae. The disease resistance-related role of RPH1 was not limited to the Arabidopsis model system. Silencing of the potato homolog StRPH1 in a resistant potato cultivar caused susceptibility to the late blight pathogen Phytophthora infestans.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Cloroplastos/metabolismo , Phytophthora/crescimento & desenvolvimento , Sequência de Aminoácidos , Arabidopsis/genética , Arabidopsis/imunologia , Proteínas de Arabidopsis/genética , Morte Celular , Cloroplastos/genética , DNA de Plantas/genética , Regulação da Expressão Gênica de Plantas , Inativação Gênica , Teste de Complementação Genética , Peróxido de Hidrogênio , Dados de Sequência Molecular , Mutagênese Insercional , Estresse Oxidativo , Doenças das Plantas/genética , Doenças das Plantas/imunologia , Alinhamento de Sequência , Análise de Sequência de DNA , Solanum tuberosum/genética , Solanum tuberosum/metabolismoAssuntos
Fungos/genética , Fungos/patogenicidade , Phytophthora/genética , Phytophthora/patogenicidade , Doenças das Plantas/microbiologia , Proteínas de Algas/genética , Sequência de Aminoácidos , Análise por Conglomerados , Códon/genética , DNA de Algas/genética , DNA Complementar/genética , DNA Fúngico/genética , Evolução Molecular , Etiquetas de Sequências Expressas , Fungos/metabolismo , Biblioteca Gênica , Lisina/biossíntese , Dados de Sequência Molecular , Filogenia , Phytophthora/metabolismo , Polissacarídeo-Liases/genética , Homologia de Sequência de Aminoácidos , Especificidade da Espécie , Virulência/genéticaRESUMO
Glutathione S-transferases (GSTs) appear to be ubiquitous in plants and have defined roles in herbicide detoxification. In contrast, little is known about their roles in normal plant physiology and during responses to biotic and abiotic stress. Forty-seven members of the GST super-family were identified in the Arabidopsis genome, grouped into four classes, with amino acid sequence identity between classes being below 25%. The two small zeta (GSTZ) and theta (GSTT) classes have related GSTs in animals while the large phi (GSTF) and tau (GSTU) classes are plant specific. As a first step to functionally characterize this diverse super-family, 10 cDNAs representing all GST classes were cloned by RT-PCR and used to study AtGST expression in response to treatment with phytohormones, herbicides, oxidative stress and inoculation with virulent and avirulent strains of the downy mildew pathogen Peronospora parasitica. The abundance of transcripts encoding AtGSTF9, AtGSTF10, AtGSTU5, AtGSTU13 and AtGSTT1 were unaffected by any of the treatments. In contrast, AtGSTF6 was upregulated by all treatments while AtGSTF2, AtGSTF8, AtGSTU19 and AtGSTZ1 each showed a selective spectrum of inducibility to the different stresses indicating that regulation of gene expression in this super-family is controlled by multiple mechanisms. The respective cDNAs were over expressed in E. coli. All GSTs except AtGSTF10 formed soluble proteins which catalysed a specific range of glutathione conjugation or glutathione peroxidase activities. Our results give further insights into the complex regulation and enzymic functions of this plant gene super-family.