Fungal plant pathogens are persistent and global food security threats. To invade their hosts they often form highly specialized infection structures, known as appressoria. The cAMP/ PKA- and MAP kinase-signaling cascades have been functionally delineated as positive-acting pathways required for appressorium development. Negative-acting regulatory pathways that block appressorial development are not known. Here, we present the first detailed evidence that the conserved Target of Rapamycin (TOR) signaling pathway is a powerful inhibitor of appressorium formation by the rice blast fungus Magnaporthe oryzae. We determined TOR signaling was activated in an M. oryzae mutant strain lacking a functional copy of the GATA transcription factor-encoding gene ASD4. Δasd4 mutant strains could not form appressoria and expressed GLN1, a glutamine synthetase-encoding orthologue silenced in wild type. Inappropriate expression of GLN1 increased the intracellular steady-state levels of glutamine in Δasd4 mutant strains during axenic growth when compared to wild type. Deleting GLN1 lowered glutamine levels and promoted appressorium formation by Δasd4 strains. Furthermore, glutamine is an agonist of TOR. Treating Δasd4 mutant strains with the specific TOR kinase inhibitor rapamycin restored appressorium development. Rapamycin was also shown to induce appressorium formation by wild type and Δcpka mutant strains on non-inductive hydrophilic surfaces but had no effect on the MAP kinase mutant Δpmk1. When taken together, we implicate Asd4 in regulating intracellular glutamine levels in order to modulate TOR inhibition of appressorium formation downstream of cPKA. This study thus provides novel insight into the metabolic mechanisms that underpin the highly regulated process of appressorium development.

Understanding how microorganisms manipulate plant innate immunity and colonize host cells is a major goal of plant pathology. Here, we report that the fungal nitrooxidative stress response suppresses host defenses to facilitate the growth and development of the important rice pathogen Magnaporthe oryzae in leaf cells. Nitronate monooxygenases encoded by NMO genes catalyze the oxidative denitrification of nitroalkanes. We show that the M. oryzae NMO2 gene is required for mitigating damaging lipid nitration under nitrooxidative stress conditions and, consequently, for using nitrate and nitrite as nitrogen sources. On plants, the Δnmo2 mutant strain penetrated host cuticles like wild type, but invasive hyphal growth in rice cells was restricted and elicited plant immune responses that included the formation of cellular deposits and a host reactive oxygen species burst. Development of the M. oryzae effector-secreting biotrophic interfacial complex (BIC) was misregulated in the Δnmo2 mutant. Inhibiting or quenching host reactive oxygen species suppressed rice innate immune responses and allowed the Δnmo2 mutant to grow and develop normally in infected cells. NMO2 is thus essential for mitigating nitrooxidative cellular damage and, in rice cells, maintaining redox balance to avoid triggering plant defenses that impact M. oryzae growth and BIC development.Global rice yields are significantly and negatively impacted each year by blast disease caused by the hemibiotrophic fungus Magnaporthe oryzae 1-3 (synonym of Pyricularia oryzae). Defining the full spectrum of molecular pro- Marroquin-Guzman et al. in Nature Microbiology 2 (2017) 2 cesses used by M. oryzae to manipulate rice innate immunity and allow fungal colonization of host cells might reveal additional sources of pathogen resistance and improve crop health. M. oryzae infects hosts by first forming specialized infection structures, appressoria, at the tips of germ tubes emerging from spores adhered to the leaf surface. 4,5 A thin penetration peg emerging from an unmelanized patch on the base of the appressorium 6 is forced through the rice leaf cuticle under hydrostatic turgor pressure 1 . In the first penetrated cell, the peg differentiates into primary hyphae then bulbous invasive hyphae (IH) that are surrounded by the plant-derived extra-invasive hyphal membrane (EIHM). Branching IH fill the first invaded cell before spreading into neighboring living rice cells at around 44 h post-inoculation. 7,8 This biotrophic growth phase progresses for 4-5 days before M. oryzae enters its necrotrophic phase.To colonize rice cells, M. oryzae must first suppress or avoid triggering two types of plant innate immunity that protect against microbial attack [9][10][11] : pathogen-associated molecular pattern (PAMP) triggered immunity (PTI), which can be suppressed by microbial effectors, and effector-triggered immunity (ETI), if effectors are detected. The biotrophic interfacial complex (BIC), a host membrane-derived structure, is formed behind M. oryzae IH in each in...

SummaryCrop destruction by the hemibiotrophic rice pathogen Magnaporthe oryzae requires plant defence suppression to facilitate extensive biotrophic growth in host cells before the onset of necrosis. How this is achieved at the genetic level is not well understood. Here, we report that a M. oryzae sirtuin, MoSir2, plays an essential role in rice defence suppression and colonization by controlling superoxide dismutase (

The conserved target of rapamycin (TOR) pathway integrates growth and development with available nutrients, but how cellular glucose controls TOR function and signaling is poorly understood. Here, we provide functional evidence from the devastating rice blast fungus Magnaporthe oryzae that glucose can mediate TOR activity via the product of a novel carbon-responsive gene, ABL1, in order to tune cell cycle progression during infection-related development. Under nutrient-free conditions, wild type (WT) M. oryzae strains form terminal plant-infecting cells (appressoria) at the tips of germ tubes emerging from three-celled spores (conidia). WT appressorial development is accompanied by one round of mitosis followed by autophagic cell death of the conidium. In contrast, Δabl1 mutant strains undergo multiple rounds of accelerated mitosis in elongated germ tubes, produce few appressoria, and are abolished for autophagy. Treating WT spores with glucose or 2-deoxyglucose phenocopied Δabl1. Inactivating TOR in Δabl1 mutants or glucose-treated WT strains restored appressorium formation by promoting mitotic arrest at G1/G0 via an appressorium- and autophagy-inducing cell cycle delay at G2/M. Collectively, this work uncovers a novel glucose-ABL1-TOR signaling axis and shows it engages two metabolic checkpoints in order to modulate cell cycle tuning and mediate terminal appressorial cell differentiation. We thus provide new molecular insights into TOR regulation and cell development in response to glucose.

The blast fungus Magnaporthe oryzae threatens global food security through the widespread destruction of cultivated rice. Foliar infection requires a specialized cell called an appressorium that generates turgor to force a thin penetration hypha through the rice cuticle and into the underlying epidermal cells, where the fungus grows for the first days of infection as a symptomless biotroph. Understanding what controls biotrophic growth could open new avenues for developing sustainable blast intervention programs. Here, using molecular genetics and live-cell imaging, we dismantled M. oryzae glucose-metabolizing pathways to reveal that the transketolase enzyme, encoded by TKL1, plays an essential role in facilitating host colonization during rice blast disease. In the absence of transketolase, Δtkl1 mutant strains formed functional appressoria that penetrated rice cuticles successfully and developed invasive hyphae (IH) in rice cells from primary hyphae. However, Δtkl1 could not undertake sustained biotrophic growth or cell-to-cell movement. Transcript data and observations using fluorescently labeled histone H1:RFP fusion proteins indicated Δtkl1 mutant strains were alive in host cells but were delayed in mitosis. Mitotic delay could be reversed and IH growth restored by the addition of exogenous ATP, a metabolite depleted in Δtkl1 mutant strains. We show that ATP might act via the TOR signaling pathway, and TOR is likely a downstream target of activation for TKL1. TKL1 is also involved in controlling the migration of appressorial nuclei into primary hyphae in host cells. When taken together, our results indicate transketolase has a novel role in mediating - via ATP and TOR signaling - an in planta-specific metabolic checkpoint that controls nuclear migration from appressoria into primary hyphae, prevents mitotic delay in early IH and promotes biotrophic growth. This work thus provides new information about the metabolic strategies employed by M. oryzae to enable rice cell colonization.

Foliar fungal pathogens challenge global food security, but how they optimize growth and development during infection is understudied. Despite adopting several lifestyles to facilitate nutrient acquisition from colonized cells, little is known about the genetic underpinnings governing pathogen adaption to host-derived nutrients. Homologs of common global and pathway-specific gene regulatory elements are likely to be involved, but their contribution to pathogenicity, and how they are connected to broader genetic networks, is largely unspecified. Here, we focus on carbon and nitrogen metabolism in foliar pathogens and consider what is known, and what is not known, about fungal exploitation of host nutrient and ask how common metabolic regulators have been co-opted to the plant-pathogenic lifestyle as well as how nutrients are utilized to drive infection.

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Cover crops (CC) have been explored in corn (Zea mays L.), cotton (Gossypium hirsutum L.), soybean [Glycine max (L.) Merr.], and wheat (Triticum aestivum L.) systems for their allelopathic potential to control weeds. However, allelopathic compounds may negatively affect these row crops by reducing germination, emergence, and grain yields. We reviewed studies that document allelopathic effects of CC on subsequent row crops in field and laboratory settings. We summarize the influence of CC management, including biomass production, planting and termination timing on allelochemical quantity. Our review found few studies documenting allelopathic effects of CC on row crops in field settings. Studies that focus on understanding yield impacts of CC on row crops should be designed to include allelopathic CC–row crop interactions. Understanding the link between CC management and allelopathic dynamics can help avoid impacts on the growth and productivity of the subsequent row crop.