Understanding the genetic pathways that regulate how pathogenic fungi respond to their environment is paramount to developing effective mitigation strategies against disease. Carbon catabolite repression (CCR) is a global regulatory mechanism found in a wide range of microbial organisms that ensures the preferential utilization of glucose over less favourable carbon sources, but little is known about the components of CCR in filamentous fungi. Here we report three new mediators of CCR in the devastating rice blast fungus Magnaporthe oryzae: the sugar sensor Tps1, the Nmr1-3 inhibitor proteins, and the multidrug and toxin extrusion (MATE)–family pump, Mdt1. Using simple plate tests coupled with transcriptional analysis, we show that Tps1, in response to glucose-6-phosphate sensing, triggers CCR via the inactivation of Nmr1-3. In addition, by dissecting the CCR pathway using Agrobacterium tumefaciens-mediated mutagenesis, we also show that Mdt1 is an additional and previously unknown regulator of glucose metabolism. Mdt1 regulates glucose assimilation downstream of Tps1 and is necessary for nutrient utilization, sporulation, and pathogenicity. This is the first functional characterization of a MATE–family protein in filamentous fungi and the first description of a MATE protein in genetic regulation or plant pathogenicity. Perturbing CCR in Δtps1 and MDT1 disruption strains thus results in physiological defects that impact pathogenesis, possibly through the early expression of cell wall–degrading enzymes. Taken together, the importance of discovering three new regulators of carbon metabolism lies in understanding how M. oryzae and other pathogenic fungi respond to nutrient availability and control development during infection.

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...

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Fungal diseases cause enormous crop losses, but defining the nutrient conditions encountered by the pathogen remains elusive. Here, we generated a mutant strain of the devastating rice pathogen Magnaporthe oryzae impaired for de novo methionine biosynthesis. The resulting methionine-requiring strain grew strongly on synthetic minimal media supplemented with methionine, aspartate or complex mixtures of partially digested proteins, but could not establish disease in rice leaves. Live-cell-imaging showed the mutant could produce normal appressoria and enter host cells but failed to develop, indicating the availability or accessibility of aspartate and methionine is limited in the plant. This is the first report to demonstrate the utility of combining biochemical genetics, plate growth tests and live-cell-imaging to indicate what nutrients might not be readily available to the fungal pathogen in rice host cells.

Increasing incidences of human disease, crop destruction and ecosystem perturbations are attributable to fungi and threaten socioeconomic progress and food security on a global scale. The blast fungus Magnaporthe oryzae is the most devastating pathogen of cultivated rice, but its metabolic requirements in the host are unclear. Here we report that a purine-requiring mutant of M. oryzae could develop functional appressoria, penetrate host cells and undergo the morphogenetic transition to elaborate bulbous invasive hyphae from primary hyphae, but further in planta growth was aborted. Invasive hyphal growth following rice cell ingress is thus dependent on de novo purine biosynthesis by the pathogen and, moreover, plant sources of purines are neither available to the mutant nor required by the wild type during the early biotrophic phase of infection. This work provides new knowledge about the metabolic interface between fungus and host that might be applicable to other important intracellular fungal pathogens.