Non-host resistance to penetration and hyphal growth of Magnaporthe oryzae in Arabidopsis.
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ABSTRACT:
SUBMITTER: Nakao M
PROVIDER: S-EPMC3240950 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Magnaporthe oryzae causes rice blast disease, but little is known about the dynamic restructuring of the actin cytoskeleton during its polarized tip growth and pathogenesis. Here, we used super-resolution live-cell imaging to investigate the dynamic organization of the actin cytoskeleton in M. oryzae during hyphal tip growth and pathogenesis. We observed a dense actin network at the apical region of the hyphae and actin filaments originating from the Spitzenkörper (Spk, the organizing center for hyphal growth and development) that formed branched actin bundles radiating to the cell membrane. The actin cross-linking protein Fimbrin (MoFim1) helps organize this actin distribution. MoFim1 localizes to the actin at the subapical collar, the actin bundles, and actin at the Spk. Knockout of MoFim1 resulted in impaired Spk maintenance and reduced actin bundle formation, preventing polar growth, vesicle transport, and the expansion of hyphae in plant cells. Finally, transgenic rice (Oryza sativa) expressing RNA hairpins targeting MoFim1 exhibited improved resistance to M. oryzae infection, indicating that MoFim1 represents an excellent candidate for M. oryzae control. These results reveal the dynamics of actin assembly in M. oryzae during hyphal tip development and pathogenesis, and they suggest a mechanism in which MoFim1 organizes such actin networks.
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Project description:Actin motor myosin proteins are the driving forces behind the active transport of vesicles, and more than 20 classes of myosin have been found to contribute to a wide range of cellular processes, including endocytosis and exocytosis, autophagy, cytokinesis and the actin cytoskeleton. In Saccharomyces cerevisiae, class V myosin Myo2 (ScMyo2p) is important for the transport of distinct sets of cargo to regions of the cell along the cytoskeleton for polarized growth. To study whether myosins play a role in the formation or function of the appressorium (infectious structure) of the rice blast fungus Magnaporthe oryzae, we identified MoMyo5 as an orthologue of ScMyo2p and characterized its function. Targeted gene disruption revealed that MoMyo5 is required for intracellular transport and is essential for hyphal growth and asexual reproduction. Although the ΔMomyo5 mutant could form appressorium-like structures, the structures were unable to penetrate host cells and were therefore non-pathogenic. We further found that MoMyo5 moves dynamically from the cytoplasm to the hyphal tip, where it interacts with MoSec4, a Rab GTPase involved in secretory transport, hyphal growth and fungal pathogenicity. Our studies indicate that class V myosin and its translocation are tightly coupled with hyphal growth, asexual reproduction, appressorium function and pathogenicity in the rice blast fungus.
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Project description:In the rice blast fungus Magnaporthe oryzae, the cAMP-PKA pathway regulates surface recognition, appressorium turgor generation, and invasive growth. However, deletion of CPKA failed to block appressorium formation and responses to exogenous cAMP. In this study, we generated and characterized the cpk2 and cpkA cpk2 mutants and spontaneous suppressors of cpkA cpk2 in M. oryzae. Our results demonstrate that CPKA and CPK2 have specific and overlapping functions, and PKA activity is essential for appressorium formation and plant infection. Unlike the single mutants, the cpkA cpk2 mutant was significantly reduced in growth and rarely produced conidia. It failed to form appressoria although the intracellular cAMP level and phosphorylation of Pmk1 MAP kinase were increased. The double mutant also was defective in plant penetration and Mps1 activation. Interestingly, it often produced fast-growing spontaneous suppressors that formed appressoria but were still non-pathogenic. Two suppressor strains of cpkA cpk2 had deletion and insertion mutations in the MoSFL1 transcription factor gene. Deletion of MoSFL1 or its C-terminal 93-aa (MoSFL1ΔCT) was confirmed to suppress the defects of cpkA cpk2 in hyphal growth but not appressorium formation or pathogenesis. We also isolated 30 spontaneous suppressors of the cpkA cpk2 mutant in Fusarium graminearum and identified mutations in 29 of them in FgSFL1. Affinity purification and co-IP assays showed that this C-terminal region of MoSfl1 was essential for its interaction with the conserved Cyc8-Tup1 transcriptional co-repressor, which was reduced by cAMP treatment. Furthermore, the S211D mutation at the conserved PKA-phosphorylation site in MoSFL1 partially suppressed the defects of cpkA cpk2. Overall, our results indicate that PKA activity is essential for appressorium formation and proper activation of Pmk1 or Mps1 in M. oryzae, and phosphorylation of MoSfl1 by PKA relieves its interaction with the Cyc8-Tup1 co-repressor and suppression of genes important for hyphal growth.
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Project description:Members of the Wheat-Induced Resistance 1 (TaWIR1) gene family are highly induced in response to a wide range of pathogens. Homologues have been identified in barley, but not in Brachypodium, whereas, in rice, only distant WIR1 candidates are known. Phylogenetic analysis placed TaWIR1a and TaWIR1b within a distinct clade of wheat transcripts, whereas TaWIR1c clustered with HvWIR1 genes. Transcripts of all three TaWIR1 genes were strongly induced by a wheat-adapted isolate of Magnaporthe oryzae. Virus-induced gene silencing of the TaWIR1 gene family had no effect on the initial penetration of epidermal cells by M. oryzae. However, following the establishment of an infection site, the fungus was able to grow more extensively within the leaf tissue, relative to control leaves, indicating a role for the TaWIR1 gene family in the cell-to-cell movement of M. oryzae. In contrast, the silencing of TaWIR1 transcripts had no effect on epidermal cell penetration by a wheat-adapted isolate of Blumeria graminis, or on the subsequent growth of hyphae. Differential transcription of TaWIR1 genes was also seen in epidermal peels, relative to the remaining leaf tissue, following inoculation with M. oryzae.
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Project description:BackgroundThe annual yield losses caused by the Rice Blast Fungus, Magnaporthe oryzae, range to the equivalent for feeding 60 million people. To ward off infection by this fungus, rice has evolved a generic basal immunity (so called compatible interaction), which acts in concert with strain-specific defence (so-called incompatible interaction). The plant-defence hormone jasmonic acid (JA) promotes the resistance to M. oryzae, but the underlying mechanisms remain elusive. To get more insight into this open question, we employ the JA-deficient mutants, cpm2 and hebiba, and dissect the JA-dependent defence signalling in rice for both, compatible and incompatible interactions.ResultsWe observe that both JA-deficient mutants are more susceptible to M. oryzae as compared to their wild-type background, which holds true for both types of interactions as verified by cytological staining. Secondly, we observe that transcripts for JA biosynthesis (OsAOS2 and OsOPR7), JA signalling (OsJAZ8, OsJAZ9, OsJAZ11 and OsJAZ13), JA-dependent phytoalexin synthesis (OsNOMT), and JA-regulated defence-related genes, such as OsBBTI2 and OsPR1a, accumulate after fungal infection in a pattern that correlates with the amplitude of resistance. Thirdly, induction of defence transcripts is weaker during compatible interaction.ConclusionThe study demonstrates the pivotal role of JA in basal immunity of rice in the resistance to M. oryzae in both, compatible and incompatible interactions.
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Project description:Rice blast caused by Magnaporthe oryzae (M. oryzae) is a major threat to global rice production. In recent years, small interference RNAs (siRNAs) and host-induced gene silencing (HIGS) has been shown to be new strategies for the development of transgenic plants to control fungal diseases and proved a useful tool to study gene function in pathogens. We here tested whether in vitro feeding artificial siRNAs (asiRNAs) could compromise M. oryzae virulence and in vivo HIGS technique could improve rice blast resistance. Our data revealed that silencing of M. oryzae MoAP1 by feeding asiRNAs targeting MoAP1 (i.e., asiR1245, asiR1362, and asiR1115) resulted in inhibited fungal growth, abnormal spores, and decreased pathogenicity. Among the asiRNAs, asiR1115 was the most inhibitory toward the rice blast fungus. Conversely, the asiRNAs targeting three other genes (i.e., MoSSADH, MoACT, and MoSOM1) had no effect on fungal growth. Transgenic rice plants expressing RNA hairpins targeting MoAP1 exhibited improved resistance to 11 tested M. oryzae strains. Confocal microscopy also revealed profoundly restricted appressoria and mycelia in rice blast-infected transgenic rice plants. Our results demonstrate that in vitro asiRNA and in vivo HIGS were useful protection approaches that may be valuable to enhance rice blast resistance.
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Project description:Host shifting and host expansion of fungal plant pathogens increases the rate of emergence of new pathogens and the incidence of disease in various crops, which threaten global food security. Magnaporthe species cause serious disease in rice, namely rice blast disease, as well as in many alternative hosts, including wheat, barley, and millet. A severe outbreak of wheat blast due to Magnaporthe oryzae occurred recently in Bangladesh, after the fungus was introduced from South America, causing great loss of yield. This outbreak of wheat blast is of growing concern, because it might spread to adjacent wheat-producing areas. Therefore, it is important to understand the host range and population structure of M. oryzae and related species for determining the evolutionary relationships among Magnaporthe species and for managing blast disease in the field. Here, we collected isolates of M. oryzae and related species from various Poaceae species, including crops and weeds surrounding rice fields, in Korea and determined their phylogenetic relationships and host species specificity. Internal transcribed spacer-mediated phylogenetic analysis revealed that M. oryzae and related species are classified into four groups primarily including isolates from rice, crabgrass, millet and tall fescue. Based on pathogenicity assays, M. oryzae and related species can infect different Poaceae hosts and move among hosts, suggesting the potential for host shifting and host expansion in nature. These results provide important information on the diversification of M. oryzae and related species with a broad range of Poaceae as hosts in crop fields.
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Project description:The exocyst, an evolutionarily conserved octameric protein complex, mediates tethering of vesicles to the plasma membrane in the early stage of exocytosis. Arabidopsis Exo70, a subunit of the exocyst complex, has been found to be involved in plant immunity. Here, we characterize the function of OsExo70B1 in rice. OsExo70B1 mainly expresses in leaf and shoot and its expression is induced by pathogen-associated molecular patterns (PAMPs) and rice blast fungus Magnaporthe oryzae (M. oryzae). Knocking out OsExo70B1 results in significantly decreased resistance and defense responses to M. oryzae compared to the wild type, including more disease lesions and enhanced fungal growth, downregulated expression of pathogenesis-related (PR) genes, and decreased reactive oxygen species accumulation. In contrast, the exo70B1 mutant does not show any defects in growth and development. Furthermore, OsExo70B1 can interact with the receptor-like kinase OsCERK1, an essential component for chitin reception in rice. Taken together, our data demonstrate that OsExo70B1 functions as an important regulator in rice immunity.
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Project description:In plant-pathogen interactions, a proper light environment affects the establishment of defense responses in plants. In our previous experiments, we found that nonhost resistance (NHR) to Pyricularia oryzae Cav. in Arabidopsis thaliana (L.) Heynh. (Arabidopsis), in diurnal conditions, varies with the inoculation time. Moreover, we indicated that the circadian clock plays an important role in regulating time-of-day differences in NHR to P. oryzae in Arabidopsis. However, the involvement of photoperiod in regulating NHR was still not understood. To determine the photoperiod role, we performed the experiments in continuous light and darkness during the early Arabidopsis-P. oryzae interaction. We found that the light period after the inoculation in the evening enhanced the resistance to penetration. However, the dark period after the inoculation in the morning suppressed the penetration resistance. Furthermore, the genetic analysis indicated that jasmonic acid, reactive oxygen species, and tryptophan-derived metabolite(s) contribute to the photoperiod regulation of NHR in Arabidopsis. The present results denote that photoperiod plays an important role in regulating time-of-day differences in NHR to P. oryzae in Arabidopsis.
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Project description:ALD1 (ABERRANT GROWTH AND DEATH2 [AGD2]-LIKE DEFENSE1) is one of the key defense regulators in Arabidopsis thaliana and Nicotiana benthamiana. In these model plants, ALD1 is responsible for triggering basal defense response and systemic resistance against bacterial infection. As well ALD1 is involved in the production of pipecolic acid and an unidentified compound(s) for systemic resistance and priming syndrome, respectively. These previous studies proposed that ALD1 is a potential candidate for developing genetically modified (GM) plants that may be resistant to pathogen infection. Here we introduce a role of ALD1-LIKE gene of Oryza sativa, named as OsALD1, during plant immunity. OsALD1 mRNA was strongly transcribed in the infected leaves of rice plants by Magnaporthe oryzae, the rice blast fungus. OsALD1 proteins predominantly localized at the chloroplast in the plant cells. GM rice plants over-expressing OsALD1 were resistant to the fungal infection. The stable expression of OsALD1 also triggered strong mRNA expression of PATHOGENESIS-RELATED PROTEIN1 genes in the leaves of rice plants during infection. Taken together, we conclude that OsALD1 plays a role in disease resistance response of rice against the infection with rice blast fungus.
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