Inhibitor of cardiolipin biosynthesis-related enzyme MoGep4 confers broad-spectrum anti-fungal activity.

Plant pathogens cause devastating diseases, leading to serious losses to agriculture. Mechanistic understanding of pathogenesis of plant pathogens lays the foundation for the development of fungicides for disease control. Mitophagy, a specific form of autophagy, is important for fungal virulence. The role of cardiolipin, mitochondrial signature phospholipid, in mitophagy and pathogenesis is largely unknown in plant pathogenic fungi. The functions of enzymes involved in cardiolipin biosynthesis and relevant inhibitors were assessed using a set of assays, including genetic deletion, plant infection, lipidomics, chemical-protein interaction, chemical inhibition, and field trials. Our results showed that the cardiolipin biosynthesis-related gene MoGEP4 of the rice blast fungus Magnaporthe oryzae regulates growth, conidiation, cardiolipin biosynthesis, and virulence. Mechanistically, MoGep4 regulated mitophagy and Mps1-MAPK phosphorylation, which are required for virulence. Chemical alexidine dihydrochloride (AXD) inhibited the enzyme activity of MoGep4, cardiolipin biosynthesis and mitophagy. Importantly, AXD efficiently inhibited the growth of 10 plant pathogens and controlled rice blast and Fusarium head blight in the field. Our study demonstrated that MoGep4 regulates mitophagy, Mps1 phosphorylation and pathogenesis in M. oryzae. In addition, we found that the MoGep4 inhibitor, AXD, displays broad-spectrum antifungal activity and is a promising candidate for fungicide development.

Rmg10, a Novel Wheat Blast Resistance Gene Derived from Aegilops tauschii.

Wheat blast, caused by Pyricularia oryzae (syn. Magnaporthe oryzae) pathotype Triticum (MoT), is a devastating disease that can result in up to 100% yield loss in affected fields. To find new resistance genes against wheat blast, we screened 199 accessions of Aegilops tauschii, the D genome progenitor of common wheat (Triticum aestivum), by seedling inoculation assays with Brazilian MoT isolate Br48 and found 14 resistant accessions. A synthetic hexaploid wheat line (Ldn/KU-2097) derived from a cross between the T. turgidum 'Langdon' (Ldn) and resistant A. tauschii accession KU-2097 exhibited resistance in seedlings and spikes against Br48. In an F2 population derived from 'Chinese Spring' × Ldn/KU-2097, resistant and susceptible individuals segregated in a 3:1 ratio, suggesting that the resistance from KU-2097 is controlled by a single dominant gene. We designated this gene Rmg10. Genetic mapping using an F2:3 population from the same cross mapped the RMG10 locus to the short arm of chromosome 2D. Rmg10 was ineffective against Bangladesh isolates but effective against Brazilian isolates. Field tests in Bolivia showed increased spike resistance in a synthetic octaploid wheat line produced from a cross between common wheat cultivar 'Gladius' and KU-2097. These results suggest that Rmg10 would be beneficial in farmers' fields in South America.

Identification of Rmg11 in Tetraploid Wheat as a New Blast Resistance Gene with Tolerance to High Temperature.

Wheat blast caused by Pyricularia oryzae pathotype Triticum has spread to Asia (Bangladesh) and Africa (Zambia) from the endemic region of South America. Wheat varieties with durable resistance are needed, but very limited resistance resources are currently available. After screening tetraploid wheat accessions, we found an exceptional accession St19 (Triticum dicoccum, KU-114). Primary leaves of St19 were resistant not only to Brazilian isolate Br48 (a carrier of Type eI of AVR-Rmg8) but also to Br48ΔA8, an AVR-Rmg8 disruptant of Br48, even at 30°C, suggesting that the resistance of St19 is tolerant to high temperature and controlled by a gene or genes other than Rmg8. When an F2 population derived from a cross between St19 and St30 (a susceptible accession of T. paleocolchicum, KU-191) was inoculated with Br48, resistant and susceptible seedlings segregated in a 3:1 ratio, indicating that resistance of St19 is conferred by a single gene. We designated this gene Rmg11. Molecular mapping revealed that the RMG11 locus is located on the short arm of chromosome 7A. Rmg11 is effective not only against other two Brazilian isolates (Br5 and Br116.5) but also against Bangladeshi isolates (T-108 and T-109) at the seedling stage. At the heading stage, lines containing Rmg11 were highly susceptible to the Bangladeshi isolates but moderately resistant to the Brazilian isolates. Stacking of Rmg11 with Rmg8 and the 2NS segment is highly recommended to achieve durable wheat blast resistance.

Breeding of a Near-Isogenic Wheat Line Resistant to Wheat Blast at Both Seedling and Heading Stages Through Incorporation of Rmg8.

Wheat blast caused by Pyricularia oryzae pathotype Triticum (MoT) has been transmitted from South America to Bangladesh and Zambia and is now spreading in these countries. To prepare against its further spread to Asian countries, we introduced Rmg8, a gene for resistance to wheat blast, into a Japanese elite cultivar, Chikugoizumi (ChI), through recurrent backcrosses and established ChI near-isogenic lines, #2-1-10 with the Rmg8/Rmg8 genotype and #4-2-10 with the rmg8/rmg8 genotype. A molecular analysis suggested that at least 96.6% of the #2-1-10 genome was derived from the recurrent parent ChI. The #2-1-10 line was resistant to MoT not only in primary leaves at the seedling stage but also in spikes and flag leaves at the heading stage. The strength of the resistance in spikes of this Rmg8 carrier was comparable to that of a carrier of the 2NS segment, which has been the only genetic resource released to farmers' fields for wheat blast resistance. On the other hand, the 2NS resistance was not expressed on leaves at the seedling stage nor flag leaves at the heading stage. Considering that leaf blast has been increasingly reported and regarded as an important inoculum source for spike blast, Rmg8 expressed at both the seedling and heading stages, or more strictly in both leaves and spikes, is suggested to be useful to prevent the spread of MoT in Asia and Africa.

A Rapid, Equipment-Free Method for Detecting Avirulence Genes of Pyricularia oryzae Using a Lateral Flow Strip-Based RPA Assay.

Rice blast, caused by Pyricularia oryzae, is one of the most destructive rice diseases worldwide. Using resistant rice varieties is the most cost-effective way to control rice blast. Consequently, it is critical to monitor the distribution frequency of avirulence (Avr) genes in rice planting fields to facilitate the breeding of resistant rice varieties. In this study, we established a rapid recombinase polymerase amplification-lateral flow dipstick (RPA-LFD) detection system for the identification of AvrPik, Avr-Piz-t, and Avr-Pi9. The optimized reaction temperature and duration were 37°C and 20 min, indicating that the reaction system could be initiated by body temperature without relying on any precision instruments. Specificity analysis showed that the primer and probe combinations targeting the three Avr genes exhibited a remarkable specificity at genus-level detection. Under the optimized condition, the lower detected thresholds of AvrPik, Avr-Piz-t, and Avr-Pi9 were 10 fg/µl, 100 fg/µl, and 10 pg/µl, respectively. Notably, the detection sensitivity of the three Avr genes was much higher than that of PCR. In addition, we also successfully detected the presence of AvrPik, Avr-Piz-t, and Avr-Pi9 in the leaf and panicle blast lesions with the RPA-LFD detection system. In particular, the genomic DNA was extracted using the simpler PEG-NaOH rapid extraction method. In summary, we developed an RPA detection system for AvrPik, Avr-Pi9, and Avr-Piz-t, combined with the PEG-NaOH rapid DNA extraction method. The innovative approach achieved rapid, real-time, and accurate detection of the three Avr genes in the field, which is helpful to understand the distribution frequency of the three Avr genes in the field and provide theoretical reference for the scientific layout of resistant rice varieties.

In Vitro and Ex Vivo Antifungal Activities of Metconazole against the Rice Blast Fungus Pyricularia oryzae.

Rice blast, caused by the filamentous fungus Pyricularia oryzae, has long been one of the major threats to almost all rice-growing areas worldwide. Metconazole, 5-(4-chlorobenzyl)-2, 2-dimethyl-1-(1H-1, 2, 4-triazol-1-ylmethyl) cyclopentanol, is a lipophilic, highly active triazole fungicide that has been applied in the control of various fungal pathogens of crops (cereals, barley, wheat), such as the Fusarium and Alternaria species. However, the antifungal activity of metconazole against P. oryzae is unknown. In this study, metconazole exhibited broad spectrum antifungal activities against seven P. oryzae strains collected from rice paddy fields and the wild type strain P131. Scanning electron microscopic analysis and fluorescein diacetate staining assays revealed that metconazole treatment damaged the cell wall integrity, cell membrane permeability and even cell viability of P. oryzae, resulting in deformed and shrunken hyphae. The supplementation of metconazole in vitro increased fungal sensitivity to different stresses, such as sodium dodecyl sulfate, congo red, sodium chloride, sorbitol and oxidative stress (H2O2). Metconazole could inhibit key virulence processes of P. oryzae, including conidial germination, germ tube elongation and appressorium formation. Furthermore, this chemical prevented P. oryzae from infecting barley epidermal cells by disturbing appressorium penetration and subsequent invasive hyphae development. Pathogenicity assays indicated a reduction of over 75% in the length of blast lesions in both barley and rice leaves when 10 µg/mL of metconazole was applied. This study provides evidence to understand the antifungal effects of metconazole against P. oryzae and demonstrates its potential in rice blast management.

Loss of PWT7, Located on a Supernumerary Chromosome, Is Associated with Parasitic Specialization of Pyricularia oryzae on Wheat.

Pyricularia oryzae, a blast fungus of gramineous plants, is composed of various host genus-specific pathotypes. The avirulence of an Avena isolate on wheat is conditioned by PWT3 and PWT4. We isolated the third avirulence gene from the Avena isolate and designated it as PWT7. PWT7 was effective as an avirulence gene only at the seedling stage or on leaves. PWT7 homologs were widely distributed in a subpopulation of the Eleusine pathotype and the Lolium pathotype but completely absent in the Triticum pathotype (the wheat blast fungus). The PWT7 homolog found in the Eleusine pathotype was one of the five genes involved in its avirulence on wheat. A comparative analysis of distribution of PWT7 and the other two genes previously identified in the Eleusine pathotype suggested that, in the course of parasitic specialization toward the wheat blast fungus, a common ancestor of the Eleusine, Lolium, Avena, and Triticum pathotypes first lost PWT6, secondly PWT7, and, finally, the function of PWT3. PWT7 or its homologs were located on core chromosomes in Setaria and Eleusine isolates but on supernumerary chromosomes in Lolium and Avena isolates. This is an example of interchromosomal translocations of effector genes between core and supernumerary chromosomes. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Population structure of Pyricularia oryzae on rice in Vietnam reveals diversified populations with four pandemic and two endemic clusters.

We characterized the genetic structure of 609 strains of Pyricularia oryzae, the fungal pathogen causing rice blast disease, in three main regions in Vietnam using microsatellites (SSR) markers. From the 447 distinct multilocus genotypes identified, six genetic clusters were defined, all of them showing elevated genetic and genotypic diversities. Four of these clusters were related to rice-attacking lineages already described at the worldwide scale, whereas the two remaining clusters were endemic to Vietnam. Strains were unevenly distributed into the six clusters depending on their groups of rice variety (indica / japonica) or type of varieties (traditional / modern) of origin, but none of the clusters was specifically related to these two factors. The highest diversity of blast population was found in Northern mountainous area, and the lowest in Red River Delta in both terms of genetic diversity and gene diversity. Hierarchical AMOVAs confirmed that all three factors considered (rice variety group, type of variety origin and geography) significantly contributed to the population structure of P. oryzae in Vietnam, with highest contribution from rice variety group. Mating types were unevenly distributed among clusters. Combined with results of female fertility and linkage disequilibirum, we hypothesized that clonal reproduction probably occurred in all clusters, but that sexual reproduction likely took place at least in some restricted areas in the Northern mountainous area for strains belonging to the cluster related to the previously described recombinant lineage (worldwide lineage 1). Our study pictures the genetic diversity, population structure and reproductive mode of the blast fungus in central and north Vietnam, and shows that the observed population structure is explained by several factors, the most important one being the variability of rice variety. All these new information might help for elaborating appropriate strategies to controlling the blast disease.

Doxorubicin inhibits phosphatidylserine decarboxylase and confers broad-spectrum antifungal activity.

As phospholipids of cell membranes, phosphatidylethanolamine (PE) and phosphatidylserine (PS) play crucial roles in glycerophospholipid metabolism. Broadly, some phospholipid biosynthesis enzymes serve as potential fungicide targets. Therefore, revealing the functions and mechanism of PE biosynthesis in plant pathogens would provide potential targets for crop disease control. We performed analyses including phenotypic characterizations, lipidomics, enzyme activity, site-directed mutagenesis, and chemical inhibition assays to study the function of PS decarboxylase-encoding gene MoPSD2 in rice blast fungus Magnaporthe oryzae. The Mopsd2 mutant was defective in development, lipid metabolism, and plant infection. The PS level increased while PE decreased in Mopsd2, consistent with the enzyme activity. Furthermore, chemical doxorubicin inhibited the enzyme activity of MoPsd2 and showed antifungal activity against 10 phytopathogenic fungi including M. oryzae and reduced disease severity of two crop diseases in the field. Three predicted doxorubicin-interacting residues are important for MoPsd2 functions. Our study demonstrates that MoPsd2 is involved in de novo PE biosynthesis and contributes to the development and plant infection of M. oryzae and that doxorubicin shows broad-spectrum antifungal activity as a fungicide candidate. The study also implicates that bacterium Streptomyces peucetius, which biosynthesizes doxorubicin, could be potentially used as an eco-friendly biocontrol agent.

Evolution of the rice blast pathogen on spatially structured rice landraces maintains multiple generalist fungal lineages.

Traditional agrosystems, where humans, crops and microbes have coevolved over long periods, can serve as models to understand the ecoevolutionary determinants of disease dynamics and help the engineering of durably resistant agrosystems. Here, we investigated the genetic and phenotypic relationship between rice (Oryza sativa) landraces and their rice blast pathogen (Pyricularia oryzae) in the traditional Yuanyang terraces of flooded rice paddies in China, where rice landraces have been grown and bred over centuries without significant disease outbreaks. Analyses of genetic subdivision revealed that indica rice plants clustered according to landrace names. Three new diverse lineages of rice blast specific to the Yuanyang terraces coexisted with lineages previously detected at the worldwide scale. Population subdivision in the pathogen population did not mirror pattern of population subdivision in the host. Measuring the pathogenicity of rice blast isolates on landraces revealed generalist life history traits. Our results suggest that the implementation of disease control strategies based on the emergence or maintenance of a generalist lifestyle in pathogens may sustainably reduce the burden of disease in crops.

The secreted immune response peptide 1 functions as a phytocytokine in rice immunity.

Small signalling peptides play important roles in various plant processes, but information regarding their involvement in plant immunity is limited. We previously identified a novel small secreted protein in rice, called immune response peptide 1 (IRP1). Here, we studied the function of IRP1 in rice immunity. Rice plants overexpressing IRP1 enhanced resistance to the virulent rice blast fungus. Application of synthetic IRP1 to rice suspension cells triggered the expression of IRP1 itself and the defence gene phenylalanine ammonia-lyase 1 (PAL1). RNA-seq results revealed that 84% of genes up-regulated by IRP1, including 13 OsWRKY transcription factors, were also induced by a microbe-associated molecular pattern (MAMP), chitin, indicating that IRP1 and chitin share a similar signalling pathway. Co-treatment with chitin and IRP1 elevated the expression level of PAL1 and OsWRKYs in an additive manner. The increased chitin concentration arrested the induction of IRP1 and PAL1 expression by IRP1, but did not affect IRP1-triggered mitogen-activated protein kinases (MAPKs) activation. Collectively, our findings indicate that IRP1 functions as a phytocytokine in rice immunity regulating MAPKs and OsWRKYs that can amplify chitin and other signalling pathways, and provide new insights into how MAMPs and phytocytokines cooperatively regulate rice immunity.

The Homeodomain-Leucine Zipper Subfamily I Contributes to Leaf Age- and Time-Dependent Resistance to Pathogens in Arabidopsis thaliana.

In Arabidopsis thaliana (Arabidopsis), nonhost resistance (NHR) is influenced by both leaf age and the moment of inoculation. While the circadian clock and photoperiod have been linked to the time-dependent regulation of NHR in Arabidopsis, the mechanism underlying leaf age-dependent NHR remains unclear. In this study, we investigated leaf age-dependent NHR to Pyricularia oryzae in Arabidopsis. Our findings revealed that this NHR type is regulated by both miR156-dependent and miR156-independent pathways. To identify the key players, we utilized rice-FOX Arabidopsis lines and identified the rice HD-Zip I OsHOX6 gene. Notably, OsHOX6 expression confers robust NHR to P. oryzae and Colletotrichum nymphaeae in Arabidopsis, with its effect being contingent upon leaf age. Moreover, we explored the role of AtHB7 and AtHB12, the Arabidopsis closest homologues of OsHOX6, by studying mutants and overexpressors in Arabidopsis-C. higginsianum interaction. AtHB7 and AtHB12 were found to contribute to both penetration resistance and post-penetration resistance to C. higginsianum in a leaf age- and time-dependent manner. These findings highlight the involvement of HD-Zip I AtHB7 and AtHB12, well-known regulators of development and abiotic stress responses, in biotic stress responses in Arabidopsis.

Long-term fertilization suppresses rice pathogens by microbial volatile compounds.

Microbial volatile organic compounds (VOCs) can suppress plant pathogens. Although fertilization strongly affects soil microbial communities, the influence of fertilization on microbial VOC-mediated suppression of pathogens has not been elucidated. Soil was sampled from a paddy field that had been subjected to the following treatments for 30 years: a no-fertilizer control, mineral fertilization (NPK), NPK combined with rice straw (NPK + S), NPK combined with chicken manure (70% NPK + 30% M). Then, within a laboratory experiment, pathogens were exposed to VOCs without physical contact to assess the impact of VOCs emitted from paddy soils on in vitro growth of the fungal rice pathogens: Pyricularia oryzae and Rhizoctonia solani. The VOCs emitted from soil reduced the mycelial biomass of P. oryzae and R. solani by 36-51% and 10-30%, respectively, compared to that of the control (no soil; no VOCs emission). Overall, the highest suppression of P. oryzae and R. solani was in the NPK and NPK + S soils, which emitted more quinones, phenols, and low alcohols than NPK + M soils. The abundances of quinones and phenols in the soil air were maximal in the NPK-fertilized soil because the low ratio of dissolved organic carbon and Olsen-P increased the population of key species such as Acidobacteriae, Anaerolineae, and Entorrhizomycetes. The abundance of alcohols was minimum in the NPK + S fertilized soil because the high SOC content decreased the population of Sordariomycetes. In conclusion, mineral fertilization affects bacterial and fungal VOC emissions, thereby suppressing the growth of R. solani and P. oryzae.

Epidermal CCA1 and PMR5 contribute to nonhost resistance in Arabidopsis.

Nonhost resistance (NHR) is the most robust and durable resistance in plants, but its spatiotemporal regulation is poorly understood. The circadian clock functions in a tissue-specific manner and regulates individual physiological processes in plants. Using mutant and RNA-seq analyses, we revealed a role of CIRCADIAN CLOCK ASSOCIATED1 (CCA1) in tissue-specific and time-of-day-specific regulation of NHR to Pyricularia oryzae (syn. Magnaporthe oryzae) in Arabidopsis thaliana (Arabidopsis). Targeted perturbation of CCA1 function in epidermis compromised time-of-day-specific regulation of NHR to P. oryzae in Arabidopsis. RNA-seq analysis showed that P. oryzae inoculation alters the transcriptome in penetration 2 (pen2) plants and identified POWDERY MILDEW RESISTANCE 5 (PMR5) as a candidate gene of direct targets of CCA1. Time-of-day-specific penetration resistance to P. oryzae was reduced in Arabidopsis pen2 pmr5 mutant plants. These findings suggest that epidermal CCA1 and PMR5 contribute to the establishment of time-of-day-specific NHR to P. oryzae in Arabidopsis.

First Report of Pyricularia oryzae Causing Blast on Wild Rice (Oryza rufipogon) in China.

Wild rice (Oryza rufipogon) is an excellent genetic resource for rice breeding programs. In June 2019, typical symptoms of blast on the leaves of wild rice cv. 'Haihong-12' were observed in a 3.3-ha field in Zhanjiang (20.93° N, 109.79° E), China. The symptoms included fusiform lesions with yellowish halo at the age of lesion, grayish-white color at the center, brown and elongated central veins at both ends of lesion, and grayish-white mold layer formed on the back of lesion under humid weather conditions. Disease incidence was more than 10%. Thirty diseased leaves were collected, and infected tissues were cut into 2 × 2 mm pieces, surface disinfected with 75% ethanol for 30 s and 2% sodium hypochlorite for 60 s and rinsed three times with sterile water. The tissues were plated onto potato dextrose agar (PDA) medium and incubated at 28 °C in darkness for 3 days. Three single-spore isolates (Pos-1, Pos-2, and Pos-3) were obtained using the method described by Jia (Jia 2009) and were subjected to further morphological and molecular identification. Colonies on PDA were light grey, with cottony mycelium. Conidiophores were solitary, erect, straight or curved, septate, and pale brown and measured 68 to 125 × 3 to 4 µm. Conidiogenous cells were sympodial and denticulate. Conidia were pale brown, pyriform, and 18.2 to 42.4 × 5.1 to 8.5 µm (n = 30) in size, with two septa. Appressoria were spherical and had the size ranging 4.3 to 6.5 × 4.7 to 6.5 µm (n = 20). These morphological features agreed with the previous description of Pyricularia oryzae Cavara (Klaubauf et al. 2014). For molecular identification, the colony PCR method with MightyAmp DNA Polymerase (Lu et al. 2012) was used to amplify the internal transcribed spacer (ITS), calmodulin (CAL), actin (ACT), -tubulin (TUB) loci of the isolates using primer pairs ITS1/ITS4, CL1C/CL2C, ACT-512F/ACT-783R, and T1/ßt2b, respectively (O'Donnell et al. 1997; Weir et al. 2012; White et al. 1990). Analysis of ITS (acc. nos. MW042176 to MW042178), ACT (MW091444 to MW091446), CAL (MW091447 to MW091449), and TUB (MW091441 to MW091443) sequences revealed 100% identity with the corresponding ITS (MH859782), ACT (MH589787), CAL (MH589663), and TUB (MH589547) sequences of P. oryzae in GenBank. A phylogenetic tree was generated based on the ITS sequences using maximum likelihood method that clustered Isolates Pos-1, Pos-2, and Pos-3 with known P. oryzae. Thus, the isolates were identified as P. oryzae. Pathogenicity tests were performed using Isolate Pos-1 in a greenhouse at 24 to 30 °C with 80% relative humidity. Individual rice plants (cv. 'Haihong-12') with three leaves were grown in 10 pots, with 50 plants per pot (40 × 60 cm). Five pots were spray inoculated with a spore suspension (105 spores/ml) until runoff from leaves, and the remaining five pots were sprayed with sterile water to serve as the controls. The test was conducted three times. Disease symptoms were observed on 10% of leaves at 10 days after inoculation, but the control plants remained healthy. The fungus was re-isolated from the diseased plants and morphologically identified as P. oryzae. Thus, this is the first report of P. oryzae causing blast on O. rufipogon in China. The results provide the information that can be used by rice breeders and fungal geneticists for further studies.

Correlation of Genomic Compartments with Contrastive Modes of Functional Losses of Host Specificity Determinants During Pathotype Differentiation in Pyricularia oryzae.

The specificity between pathotypes of Pyricularia oryzae and genera of gramineous plants is governed by gene-for-gene interactions. Here, we show that avirulence genes involved in this host specificity have undergone different modes of functional losses dependent on or affected by genomic compartments harboring them. The avirulence of an Eleusine pathotype on wheat is controlled by five genes, including PWT3, which played a key role in the evolution of the Triticum pathotype (the wheat blast fungus). We cloned another gene using an association of its presence or absence with pathotypes and designated it as PWT6. PWT6 was widely distributed in a lineage composed of Eleusine and Eragrostis isolates but was completely absent in a lineage composed of Lolium and Triticum isolates. On the other hand, PWT3 homologs were present in all isolates, and their loss of function in Triticum isolates was caused by insertions of transposable elements or nucleotide substitutions. Analyses of whole-genome sequences of representative isolates revealed that these two genes were located in different genomic compartments; PWT6 was located in a repeat-rich region, while PWT3 was located in a repeat-poor region. These results suggest that the course of differentiation of the pathotypes in P. oryzae appears to be illustrated as processes of functional losses of avirulence genes but that modes of the losses are affected by genomic compartments in which they reside.[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.

The rice RNase P protein subunit Rpp30 confers broad-spectrum resistance to fungal and bacterial pathogens.

RNase P functions either as a catalytic ribonucleoprotein (RNP) or as an RNA-free polypeptide to catalyse RNA processing, primarily tRNA 5' maturation. To the growing evidence of non-canonical roles for RNase P RNP subunits including regulation of chromatin structure and function, we add here a role for the rice RNase P Rpp30 in innate immunity. This protein (encoded by LOC_Os11g01074) was uncovered as the top hit in yeast two-hybrid assays performed with the rice histone deacetylase HDT701 as bait. We showed that HDT701 and OsRpp30 are localized to the rice nucleus, OsRpp30 expression increased post-infection by Pyricularia oryzae (syn. Magnaporthe oryzae), and OsRpp30 deacetylation coincided with HDT701 overexpression in vivo. Overexpression of OsRpp30 in transgenic rice increased expression of defence genes and generation of reactive oxygen species after pathogen-associated molecular pattern elicitor treatment, outcomes that culminated in resistance to a fungal (P. oryzae) and a bacterial (Xanthomonas oryzae pv. oryzae) pathogen. Knockout of OsRpp30 yielded the opposite phenotypes. Moreover, HA-tagged OsRpp30 co-purified with RNase P pre-tRNA cleavage activity. Interestingly, OsRpp30 is conserved in grass crops, including a near-identical C-terminal tail that is essential for HDT701 binding and defence regulation. Overall, our results suggest that OsRpp30 plays an important role in rice immune response to pathogens and provides a new approach to generate broad-spectrum disease-resistant rice cultivars.

Bayogenin 3-O-cellobioside confers non-cultivar-specific defence against the rice blast fungus Pyricularia oryzae.

Rice cultivars from japonica and indica lineage possess differential resistance against blast fungus as a result of genetic divergence. Whether different rice cultivars also show distinct metabolomic changes in response to P. oryzae, and their role in host resistance, are poorly understood. Here, we examine the responses of six different rice cultivars from japonica and indica lineage challenged with P. oryzae. Both susceptible and resistant rice cultivars expressed several metabolites exclusively during P. oryzae infection, including the saponin Bayogenin 3-O-cellobioside. Bayogenin 3-O-cellobioside level in infected rice directly correlated with their resistant attributes. These findings reveal, for the first time to our knowledge that besides oat, other grass plants including rice produces protective saponins. Our study provides insight into the role of pathogen-mediated metabolomics reprogramming in host immunity. The correlation between Bayogenin 3-O-Cellobioside levels and blast resistance suggests that engineering saponin expression in cereal crops represents attractive and sustainable disease management.

The CC-NB-LRR OsRLR1 mediates rice disease resistance through interaction with OsWRKY19.

Nucleotide-binding site-leucine-rich repeat (NB-LRR) resistance proteins are critical for plant resistance to pathogens; however, their mechanism of activation and signal transduction is still not well understood. We identified a mutation in an as yet uncharacterized rice coiled-coil (CC)-NB-LRR, Oryza sativa RPM1-like resistance gene 1 (OsRLR1), which leads to hypersensitive response (HR)-like lesions on the leaf blade and broad-range resistance to the fungal pathogen Pyricularia oryzae (syn. Magnaporthe oryzae) and the bacterial pathogen Xanthomonas oryzae pv. oryzae, together with strong growth reduction. Consistently, OsRLR1-overexpression lines showed enhanced resistance to both pathogens. Moreover, we found that OsRLR1 mediates the defence response through direct interaction in the nucleus with the transcription factor OsWRKY19. Down-regulation of OsWRKY19 in the rlr1 mutant compromised the HR-like phenotype and resistance response, and largely restored plant growth. OsWRKY19 binds to the promoter of OsPR10 to activate the defence response. Taken together, our data highlight the role of a new residue involved in the NB-LRR activation mechanism, allowing identification of a new NB-LRR downstream signalling pathway.

Suppression of wheat blast resistance by an effector of Pyricularia oryzae is counteracted by a host specificity resistance gene in wheat.

Wheat blast caused by the Triticum pathotype of Pyricularia oryzae poses a serious threat to wheat production in South America and Asia and is now becoming a pandemic disease. Here, we show that Rmg8, a promising wheat gene for resistance breeding, is suppressed by PWT4, an effector gene of P. oryzae, and in turn that the suppression is counteracted by Rwt4, a wheat gene recognizing PWT4. When PWT4 was introduced into a wheat blast isolate carrying AVR-Rmg8 (an avirulence gene corresponding to Rmg8), PWT4 suppressed wheat resistance conferred by Rmg8. PWT4 did not alter the expression of AVR-Rmg8, but higher expression of PWT4 led to more efficient suppression. This suppression was observed in rwt4 carriers, but not in Rwt4 carriers, indicating that it is counteracted by Rwt4. PWT4 was assumed to have been horizontally transferred from a weed-associated cryptic species, P. pennisetigena, to an Avena isolate of P. oryzae in Brazil. This implies a potential risk of the acquisition of PWT4 by the wheat blast fungus and the 'breakdown' of Rmg8. We suggest that Rmg8 should be introduced together with Rwt4 into a wheat cultivar when it is used for resistance breeding.