Common themes in nutrient acquisition by plant symbiotic microbes,described by the Gene Ontology

A critical function for symbionts is the acquisition of nutrients from their host. Relationships between hosts and symbionts range from biotrophic mutualism to necrotrophic parasitism, with a corresponding range of structures to facilitate nutrient flow between host and symbiont. Here, we review common themes among the nutrient acquisition strategies of a range of plant symbiotic microorganisms, including mutualistic symbionts, biotrophic pathogens that feed from living tissue, necrotrophic pathogens that kill host tissue, and hemibiotrophic pathogens that switch from biotrophy to necrotrophy. We show how Gene Ontology (GO) terms developed by the Plant-Associated Microbe Gene Ontology (PAMGO) Consortium can be used for describing commonalities in nutrient acquisition among diverse plant symbionts. Where appropriate, parallels found among animal symbionts are also highlighted.     

The biotrophy-associated secreted protein 4 (BAS4) participates in the transition of Magnaporthe oryzae from the biotrophic to the necrotrophic phase

The physiological and metabolic processes of host plants are manipulated and remodeled by phytopathogenic fungi during infection, revealed obvious signs of biotrophy of the hemibiotrophic pathogen. As we known that effector proteins play key roles in interaction of hemibiotrophic fungi and their host plants. BAS4 (biotrophy-associated secreted protein 4) is an EIHM (extrainvasive hyphal membrane) matrix protein that was highly expressed in infectious hyphae. In order to study whether BAS4 is involved in the transition of rice blast fungus from biotrophic to necrotrophic phase, The susceptible rice cultivar Lijiangxintuanheigu (LTH) that were pre-treated with prokaryotic expression product of BAS4 and then followed with inoculation of the blast strain, more serious blast disease symptom, more biomass such as sporulation and fungal relative growth, and lower expression level of pathogenicity-related genes appeared in lesion of the rice leaves than those of the PBS-pretreated-leaves followed with inoculation of the same blast strain, which demonstrating that BAS4 invitro changed rice defense system to facilitate infection of rice blast strain. And the susceptible rice cultivar (LTH) were inoculated withBAS4-overexpressed blast strain, we also found more serious blast disease symptom and more biomass also appeared in lesion of leaves inoculated with BAS4-overexpressed strain than those of leaves inoculated with the wild-type strain, and expression level of pathogenicity-related genes appeared lower in biotrophic phase and higher in necrotrophic phase of infection, indicating BAS4 maybe in vivo regulate defense system of rice to facilitate transition of biotrophic to necrotrophic phase. Our data demonstrates that BAS4 in vitro and in vivo participates in transition from the biotrophic to the necrotrophic phase of Magnaporthe oryzae.     

Mitochondrial dynamics and mitophagy are necessary for proper invasive growth in rice blast

Magnaporthe oryzae causes blast disease, which is one of the most devastating infections in rice and several important cereal crops. Magnaporthe oryzae needs to coordinate gene regulation, morphological changes, nutrient acquisition and host evasion in order to invade and proliferate within the plant tissues. Thus far, the molecular mechanisms underlying the regulation of invasive growth in planta have remained largely unknown. We identified a precise filamentous-punctate-filamentous cycle in mitochondrial morphology during Magnaporthe–rice interaction. Interestingly, disruption of such mitochondrial dynamics by deletion of genes regulating either the mitochondrial fusion (MoFzo1) or fission (MoDnm1) machinery, or inhibition of mitochondrial fission using Mdivi-1 caused significant reduction in M. oryzae pathogenicity. Furthermore, exogenous carbon source(s) but not antioxidant treatment delayed such mitochondrial dynamics/transition during invasive growth. In contrast, carbon starvation induced the breakdown of the mitochondrial network and led to more punctate mitochondria in vitro. Such nutrient-based regulation of organellar dynamics preceded MoAtg24-mediated mitophagy, which was found to be essential for proper biotrophic development and invasive growth in planta. We propose that precise mitochondrial dynamics and mitophagy occur during the transition from biotrophy to necrotrophy and are required for proper induction and establishment of the blast disease in rice.     

A nonclassically secreted effector of Magnaporthe oryzae targets host nuclei and plays important roles in fungal growth and plant infection

Rice blast caused by Magnaporthe oryzae is one of the most destructive diseases and poses a growing threat to food security worldwide. Like many other filamentous pathogens, rice blast fungus releases multiple types of effector proteins to facilitate fungal infection and modulate host defence responses. However, most of the characterized effectors contain an N-terminal signal peptide. Here, we report the results of the functional characterization of a nonclassically secreted nuclear targeting effector in M. oryzae (MoNte1). MoNte1 has no signal peptide, but can be secreted and translocated into plant nuclei driven by a nuclear targeting peptide. It could also induce hypersensitive cell death when transiently expressed in Nicotiana benthamiana. Deletion of the MoNTE1 gene caused a significant reduction of fungal growth and conidiogenesis, partially impaired appressorium formation and host colonization, and also dramatically attenuated the pathogenicity. Taken together, these findings reveal a novel effector secretion pathway and deepen our understanding of rice–M. oryzae interactions.     

Characterizing Roles for the Glutathione Reductase,Thioredoxin Reductase and Thioredoxin Peroxidase-Encoding Genes of Magnaporthe oryzae during Rice Blast Disease

Understanding how pathogenic fungi adapt to host plant cells is of major concern to securing global food production. The hemibiotrophic rice blast fungus Magnaporthe oryzae, cause of the most serious disease of cultivated rice, colonizes leaf cells asymptomatically as a biotroph for 4–5 days in susceptible rice cultivars before entering its destructive necrotrophic phase. During the biotrophic growth stage, M. oryzae remains undetected in the plant while acquiring nutrients and growing cell-to-cell. Which fungal processes facilitate in planta growth and development are still being elucidated. Here, we used gene functional analysis to show how components of the NADPH-requiring glutathione and thioredoxin antioxidation systems of M. oryzae contribute to disease. Loss of glutathione reductase, thioredoxin reductase and thioredoxin peroxidase-encoding genes resulted in strains severely attenuated in their ability to grow in rice cells and that failed to produce spreading necrotic lesions on the leaf surface. Glutathione reductase, but not thioredoxin reductase or thioredoxin peroxidase, was shown to be required for neutralizing plant generated reactive oxygen species (ROS). The thioredoxin proteins, but not glutathione reductase, were shown to contribute to cell-wall integrity. Furthermore, glutathione and thioredoxin gene expression, under axenic growth conditions, was dependent on both the presence of glucose and the M. oryzae sugar/ NADPH sensor Tps1, thereby suggesting how glucose availability, NADPH production and antioxidation might be connected. Taken together, this work identifies components of the fungal glutathione and thioredoxin antioxidation systems as determinants of rice blast disease that act to facilitate biotrophic colonization of host cells by M. oryzae.     

Two Magnaporthe appressoria‐specific (MAS) proteins,MoMas3 and MoMas5, are required for suppressing host innate immunity and promoting biotrophic growth in rice cells

In the devastating rice blast fungus Magnaporthe oryzae, six Magnaporthe appressoria‐specific (MAS) proteins are encoded by MoGAS1, MoGAS2 and MoMAS3–MoMAS6. MoGAS1 and MoGAS2 were previously characterized as M. oryzae virulence factors; however, the roles of the other four genes are unknown. Here, we found that, although the loss of any MAS gene did not affect appressorial formation or vegetative growth, ∆Momas3 and ∆Momas5 mutant strains (but not the others) were reduced in virulence on susceptible CO‐39 rice seedlings. Focusing on ∆Momas3 and ∆Momas5 mutant strains, we found that they could penetrate host leaf surfaces and fill the first infected rice cell but did not spread readily to neighbouring cells, suggesting they were impaired for biotrophic growth. Live‐cell imaging of fluorescently labelled MoMas3 and MoMas5 proteins showed that during biotrophy, MoMas3 localized to the apoplastic compartment formed between fungal invasive hyphae and the plant‐derived extra‐invasive hyphal membrane while MoMas5 localized to the appressoria and the penetration peg. The loss of either MoMAS3 or MoMAS5 resulted in the accumulation of reactive oxygen species (ROS) in infected rice cells, resulting in the triggering of plant defences that inhibited mutant growth in planta. ∆Momas3 and ∆Momas5 biotrophic growth could be remediated by inhibiting host NADPH oxidases and suppressing ROS accumulation. Thus, MoMas3 and MoMas5 are novel virulence factors involved in suppressing host plant innate immunity to promote biotrophic growth.     

Identification and characterization of suppressors of plant cell death (SPD) effectors from Magnaporthe oryzae

Phytopathogenic microorganisms, including the fungal pathogen Magnaporthe oryzae, secrete a myriad of effector proteins to facilitate infection. Utilizing the transient expression of candidate effectors in the leaves of the model plant Nicotiana benthamiana, we identified 11 suppressors of plant cell death (SPD) effectors from M. oryzae that were able to block the host cell death reaction induced by Nep1. Ten of these 11 were also able to suppress BAX‐mediated plant cell death. Five of the 11 SPD genes have been identified previously as either essential for the pathogenicity of M. oryzae, secreted into the plant during disease development, or as suppressors or homologues of other characterized suppressors. In addition, of the remaining six, we showed that SPD8 (previously identified as BAS162) was localized to the rice cytoplasm in invaded and surrounding uninvaded cells during biotrophic invasion. Sequence analysis of the 11 SPD genes across 43 re‐sequenced M. oryzae genomes revealed that SPD2, SPD4 and SPD7 have nucleotide polymorphisms amongst the isolates. SPD4 exhibited the highest level of nucleotide diversity of any currently known effector from M. oryzae in addition to the presence/absence polymorphisms, suggesting that this gene is potentially undergoing selection to avoid recognition by the host. Taken together, we have identified a series of effectors, some of which were previously unknown or whose function was unknown, that probably act at different stages of the infection process and contribute to the virulence of M. oryzae.     

When and how to kill a plant cell: Infection strategies of plant pathogenic fungi

Fungi cause severe diseases on a broad range of crop and ornamental plants, leading to significant economical losses. Plant pathogenic fungi exhibit a huge variability in their mode of infection, differentiation and function of infection structures and nutritional strategy. In this review, advances in understanding mechanisms of biotrophy, necrotrophy and hemibiotrophic lifestyles are described. Special emphasis is given to the biotrophy-necrotrophy switch of hemibiotrophic pathogens, and to biosynthesis, chemical diversity and mode of action of various fungal toxins produced during the infection process.     

Identification and characterization of Cercospora beticola necrosis-inducing effector CbNip1

Cercospora beticola is a hemibiotrophic fungus that causes cercospora leaf spot disease of sugar beet (Beta vulgaris). After an initial symptomless biotrophic phase of colonization, necrotic lesions appear on host leaves as the fungus switches to a necrotrophic lifestyle. The phytotoxic secondary metabolite cercosporin has been shown to facilitate fungal virulence for several Cercospora spp. However, because cercosporin production and subsequent cercosporin-initiated formation of reactive oxygen species is light-dependent, cell death evocation by this toxin is only fully ensured during a period of light. Here, we report the discovery of the effector protein CbNip1 secreted by C. beticola that causes enhanced necrosis in the absence of light and, therefore, may complement light-dependent necrosis formation by cercosporin. Infiltration of CbNip1 protein into sugar beet leaves revealed that darkness is essential for full CbNip1-triggered necrosis, as light exposure delayed CbNip1-triggered host cell death. Gene expression analysis during host infection shows that CbNip1 expression is correlated with symptom development in planta. Targeted gene replacement of CbNip1 leads to a significant reduction in virulence, indicating the importance of CbNip1 during colonization. Analysis of 89 C. beticola genomes revealed that CbNip1 resides in a region that recently underwent a selective sweep, suggesting selection pressure exists to maintain a beneficial variant of the gene. Taken together, CbNip1 is a crucial effector during the C. beticola–sugar beet disease process.