Molecular plant pathology最新文献

Mad2, a conserved core component of the spindle assembly checkpoint (SAC) in eukaryotes, delays anaphase onset in case of incorrect kinetochore-microtubule attachment. However, its functions in plant-pathogenic fungi remain largely unknown. Here, we identified the Mad2 homologue in rice blast fungus Magnaporthe oryzae (MoMad2), which shows high similarity with Mad2 in fission yeast. When expressed in fission yeast, MoMad2 associated with native SpMad1 and SpCdc20, and successfully rescued the ΔSpmad2 mutant's defect in arresting anaphase onset upon damaged spindle, indicating the conserved SAC function of MoMad2. Moreover, MoMad2 interacted with MoMad1 and depends on MoMad1 for its nuclear envelope-localisation. Although it plays a dispensable role in M. oryzae growth, MoMad2 is required for tolerance to the microtubule depolymerising agent treatment. ΔMomad2 mutants exhibited shorter hyphal compartments and earlier conidial germination and appressorium formation, suggesting that MoMad2 deletion shortens M. oryzae's mitotic cell cycle due to defective SAC arrest. Additionally, knockout of MoMAD2 decreased the appressorial turgor pressure, impaired appressorium penetration and compromised M. oryzae pathogenicity. Taken together, our findings revealed that MoMad2, as a conserved component in SAC signalling, is essential for full pathogenicity of rice blast fungus.

Potato mop-top virus (PMTV) is the causal agent of potato tuber spraing disease, which causes significant economic losses to potato production worldwide. The 3'-proximal end of PMTV genomic RNA3 encodes an 8 kDa cysteine-rich protein (8K) that is not essential for replication and movement but contributes to virus infection and symptom development. Here, we demonstrate that PMTV 8K forms endomembrane multimers, alters the membrane permeability of Escherichia coli, and possesses potassium and proton conductance activity. In addition, our data reveal that two conserved cysteine residues in the central hydrophobic α-helix are essential for the viroporin activity. These results not only deepen our understanding of the function of PMTV 8K but also provide new insights into the diversity and origin of plant viral viroporins.

Root rot disease in Panax notoginseng, primarily caused by the pathogenic fungus Fusarium solani, significantly impacts the growth and production of this medicinal herb. To elucidate the defence mechanisms of P. notoginseng against root rot, we employed proteomics and gas chromatography-mass spectrometry (GC-MS)-based metabolomics analyses. These analyses revealed significant accumulations of metabolites involved in phenylpropanoid, terpenoid and steroid biosynthesis pathways in F. solani-infected P. notoginseng roots. This accumulation correlated with the up-regulation of synthetases in these pathways as indicated by proteomics data. Focusing on stigmasterol, a representative steroid with differential accumulation levels, and its biosynthesis gene PnCYP710A, we investigated the role of stigmasterol metabolism in the defence response against root rot. Stigmasterol exhibited significant inhibitory effects on spore germination and hyphal growth of F. solani. Furthermore, PnCYP710A was up-regulated upon F. solani infection and induced by hormonal signals such as methyl jasmonate (MeJA). Overexpression of PnCYP710A in tobacco enhanced resistance to F. solani, up-regulated expression of JA biosynthesis/signalling pathway-related genes, increased accumulation of stigmasterol/lignin/callus, and maintained reactive oxygen species homeostasis during F. solani infection. Conversely, RNA interference (RNAi) of PnCYP710A in P. notoginseng yielded opposite effects. Additionally, PnWRKY4 positively regulated the transcription level of PnCYP710A by binding to its promoter. In summary, this study not only identifies volatile metabolites and proteins involved in the defence response of P. notoginseng against root rot but also discovers that PnWRKY4 activates stigmasterol biosynthesis to resist root rot pathogen infection.

Among eukaryotes, Rab GTPases are critical for intracellular membrane trafficking and possess various functions. Oomycetes, responsible for many devastating plant diseases, pose a significant threat to global agriculture. However, the functions of Rab GTPases in oomycetes are largely uncharted. In this study, we functionally characterised two Rab8 homologues, PlRab8A and PlRab8B, from a plant-pathogenic oomycete, Peronophythora litchii. These genes are crucial for mycelial growth and play significant roles in the production and morphology of sporangia, as well as the formation of zoospores. PlRab8A and PlRab8B also contribute to maintaining cell wall integrity, osmotic stress tolerance and oospore formation, which are vital for the organism's survival and virulence. Moreover, PlRab8A and PlRab8B are essential for the full virulence of P. litchii by influencing laccase activity. We also found that PlRab8B co-localised with lipid droplets (LDs) and negatively affected LD accumulation. Autophagy was enhanced and autophagy-related genes (PlATGs) were up-regulated in PlRab8B mutants. Additionally, PlRab8A was found to interact with PlRab6. Similar to PlRab8, PlRab6 is also required for growth, zoospore formation and pathogenicity of P. litchii. Taken together, this study reported for the first time the functions of three Rab proteins regulating autophagy, LD accumulation and pathogenesis in oomycetes.

Movement proteins (MPs) modulate the size exclusion limit of plasmodesmata-membrane-lined channels connecting plant cells-thereby allowing cell-to-cell movement and systemic spread of plant viruses. The largest and arguably best-studied group of MPs is the 30K superfamily. Its family members share little sequence similarity, with only a handful of residues being well conserved. Yet, all family members appear to adopt the same jelly-roll protein fold structure. Lettuce big-vein associated virus (LBVaV), a member of the Rhabdoviridae family, is closely associated with lettuce big-vein disease (LBVD). It appears to facilitate the long-distance movement of Mirafiori lettuce big-vein virus (MiLBVV) in plants through an unknown mechanism. Notably, enhanced MiLBVV spread correlates with severe LBVD symptoms. Despite LBVaV having been known for decades, its proteins have not been studied in detail thus far. By using a combination of Alphafold2 structure modelling and FoldSeek structure-based homology searches, we managed to annotate all LBVaV open reading frames (ORFs), with ORF3 clustering with the 30K superfamily. While ORF3 is the most conserved protein sequence among the LBVaV-encoded ORFs, it shares only 5%-11% protein sequence identity with related MPs in the same genus. Microscopy studies confirmed that ORF3 locates at plasmodesmata, and in planta expression of ORF3 allowed cell-to-cell movement of two movement-impaired plant viruses. Thus, the Alphafold2-FoldSeek strategy allowed successful annotation of a plant viral genome even when viral proteins show little sequence similarity.

Filamentous fungi have evolved compartmentalised genomes comprising conserved core regions and dynamic accessory regions, which are thought to drive adaptation to changing environments, including interactions with host organisms. Tropical Race 4 (TR4) is a lineage of banana-infecting Fusarium spp. and causes a devastating Fusarium wilt epidemic in the industrial banana cultivar Cavendish. A recent study showed that TR4 contains a single accessory chromosome (chromosome 12), which in some strains has undergone extensive internal duplications, tripling its size compared to other closely related strains. However, the contribution of this accessory chromosome to virulence is currently unknown. Here we show that the induced loss of accessory chromosome 12 in the TR4 reference strain II5 leads to reduced virulence on banana plants. Moreover, loss of chromosome 12 co-occurs with structural rearrangements of conserved core chromosomes. Together, our results provide new insights into the chromosome dynamics of the banana-infecting Fusarium TR4 lineage and highlight the importance of its unique accessory chromosome in virulence.

Superinfection exclusion (SIE) is a finely tuned virus-virus interaction mechanism closely linked to the viral infection cycle. However, the mechanistic basis of SIE remains incompletely understood in plant viruses, particularly among negative-sense, single-stranded RNA viruses. In this study, we first describe the development of an efficient reverse genetics system for the plant nucleorhabdovirus Physostegia chlorotic mottle virus (PhCMoV) by codon optimisation of the large polymerase coding sequence. Using fluorescently tagged variants of PhCMoV, as well as three additional closely or distantly related plant rhabdoviruses, we found that each rhabdovirus displayed homotypic SIE. Moreover, two closely related alphanucleorhabdoviruses, PhCMoV and eggplant mottled dwarf virus, also exhibited mutual exclusion. Loss- and gain-of-function reverse genetics analyses identified the rhabdovirus matrix (M) protein as the central SIE effector: M-deficient mutant viruses lost exclusion capacity, whereas ectopically expressed heterologous M proteins conferred SIE against otherwise compatible, distantly related rhabdoviruses. Additional functional assays demonstrated that the ability of rhabdovirus M proteins to suppress cognate and noncognate viral RNA synthesis correlated with the intra- and interspecies SIE capacity. The widespread occurrence of SIE across distinct plant rhabdoviruses underscores its importance for understanding the viral replication cycle and highlights its practical relevance for the development of novel virus control strategies.

In previous work, we have shown that the transcription factor Nit2 plays a major role in the utilisation of non-favoured nitrogen sources like nitrate, minor amino acids or nucleobases in saprotrophic sporidia of the basidiomycete corn smut fungus Ustilago maydis. Addressing the knowledge gap regarding how filamentous phytopathogens adapt to nitrogen limitation in the host plant, we employed Δnit2 mutants in the natural FB1 × FB2 background to identify Nit2-regulated genes during biotrophy. We further investigated the impact of Nit2 on the physiology of leaf galls in nitrogen-replete versus nitrogen-limited host plants by comparative RNA-Seq and metabolic steady state analysis. About one third of the fungal genes affected by Nit2 during biotrophy were involved in nitrogen metabolism and transport, only showing minor overlap to saprotrophic sporidia. Induction of the nitrate assimilation cluster was completely dependent on Nit2 during biotrophy. In nitrogen-limited host plants, Δnit2 leaf galls accumulated nitrate and showed reduced accumulation of the nitrogen-rich phloem transport amino acids asparagine and glutamine compared to wild-type galls. However, total protein content in galls and pathogenicity were comparable between fungal genotypes in both nitrogen regimes. The findings of our physiological and transcriptomic analysis demonstrate that nitrate utilisation is dispensable for U. maydis during biotrophy and can likely be actively compensated by increased utilisation of abundant organic nitrogen sources, like asparagine, GABA and glutamine in a partially Nit2-dependent fashion.

Colletotrichum graminicola, the maize anthracnose fungus, is known for its ability to invade above-ground tissues. This fungus forms two distinct asexual spore types in its life cycle, falcate conidia in acervuli on infected leaves and oval conidia in parenchyma cells in leaf and stem lesions. Our study reveals a previously unknown role for oval conidia in the infection of roots. We investigated whether root exudate from maize could redirect the growth of germlings generated by C. graminicola. Our results showed that only germlings derived from oval conidia were able to respond to root exudates, whereas those from falcate conidia did not. High-performance liquid chromatography/mass spectrometry (HPLC-MS) analyses combined with biological assays and genetic studies identified diterpenoids from maize as attractants perceived by the a-pheromone receptor CgSte3. We further explored the root-fungus interaction by analysing the germination of oval and falcate spores in soils of different composition. Oval conidia germinated rapidly under all conditions, whereas falcate conidia remained dormant and became highly vacuolated even in the presence of the host plant. Microscopic evaluation of root infection experiments showed that both conidia types attached to root material and formed penetration structures. However, only oval conidia enabled the pathogenic fungus to reach upper plant parts from infected roots. Our findings suggest a novel role for oval conidia in the infection of roots, highlighting the complexity of the anthracnose disease cycle.