Project description:BACKGROUND:Nicotiana attenuata is an ecological model plant whose 2.57 Gb genome has recently been sequenced and assembled and for which miRNAs and their genomic locations have been identified. To understand how this plant's miRNAs are reconfigured during plant-arbuscular mycorrhizal fungal (AMF) interactions and whether hostplant calcium- and calmodulin dependent protein kinase (CCaMK) expression which regulates the AMF interaction also modulates miRNAs levels and regulation, we performed a large-scale miRNA analysis of this plant-AMF interaction. RESULTS:Next generation sequencing of miRNAs in roots of empty vector (EV) N. attenuata plants and an isogenic line silenced in CCaMK expression (irCCaMK) impaired in AMF-interactions grown under competitive conditions with and without AMF inoculum revealed a total of 149 unique miRNAs: 67 conserved and 82 novel ones. The majority of the miRNAs had a length of 21 nucleotides. MiRNA abundances were highly variable ranging from 400 to more than 25,000 reads per million. The miRNA profile of irCCaMK plants impaired in AMF colonization was distinct from fully AMF-functional EV plants grown in the same pot. Six conserved miRNAs were present in all conditions and accumulated differentially depending on treatment and genotype; five (miR6153, miR403a-3p, miR7122a, miR167-5p and miR482d, but not miR399a-3p) showed the highest accumulation in AMF inoculated EV plants compared to inoculated irCCaMK plants. Furthermore, the accumulation patterns of sequence variants of selected conserved miRNAs showed a very distinct pattern related to AMF colonization - one variant of miR473-5p specifically accumulated in AMF-inoculated plants. Also abundances of miR403a-3p, miR171a-3p and one of the sequence variants of miR172a-3p increased in AMF-inoculated EV compared to inoculated irCCaMK plants and to non-inoculated EV plants, while miR399a-3p was most strongly enriched in AMF inoculated irCCaMK plants grown in competition with EV. The analysis of putative targets of selected miRNAs revealed an involvement in P starvation (miR399), phytohormone signaling (Nat-R-PN59, miR172, miR393) and defense (e.g. miR482, miR8667, Nat-R-PN-47). CONCLUSIONS:Our study demonstrates (1) a large-scale reprograming of miRNAs induced by AMF colonization and (2) that the impaired AMF signaling due to CCaMK silencing and the resulting reduced competitive ability of irCCaMK plants play a role in modulating signal-dependent miRNA accumulation.
Project description:Almost all living plants can be simultaneously colonised by arbuscular mycorrhizal fungi in the roots and endophytes in the shoots, while also being attacked by insect herbivores. However, to date, no study has ever examined the multitrophic interactions between these two different fungal groups and insects on any species of forb. Here, we examined the effects of two commercial species mixtures of arbuscular mycorrhizal fungi (AMF) and two foliar endophytes (Colletotrichum acutatum and Cladosporium oxysporum) on the growth of an invasive weed, Impatiens glandulifera, and the aphids that attack it. AMF reduced plant biomass, which was most evident when C. oxysporum was inoculated. Mycorrhizal fungi had few effects on aphids, and these depended on the identity of the endophytes present. Meanwhile, endophytes tended to increase aphid numbers, but this depended on the identity of the AMF inoculum. Throughout, there were differences in the responses of the plant to the two mycorrhizal mixtures, demonstrating clear AMF specificity in this plant. These specific effects were also strongly affected by the endophytes, with a greater number of interactions found between the AMF and endophytes than between the endophytes themselves. In particular, AMF reduced infection levels by the endophytes, while some endophyte inoculations reduced mycorrhizal colonisation. We suggest that both AMF and endophytes could play an important part in future biological control programmes of weeds, but further multitrophic experiments are required to unravel the complexity of interactions between spatially separated parts of the plant microbiome.
Project description:The virtual transcriptome of G. intraradices (based on >430,000 reads) served as the basis for the design of an EST expresson array and for a number of analyses of gene expression in germinating spores, extraradical mycelium (ERM) and symbiotic root tissues.
Project description:Roots of most terrestrial plants form symbiotic associations (mycorrhiza) with soil- borne arbuscular mycorrhizal fungi (AMF). Many studies show that mycorrhizal colonization enhances plant resistance against pathogenic fungi. However, the mechanism of mycorrhiza-induced disease resistance remains equivocal. In this study, we found that mycorrhizal inoculation with AMF Funneliformis mosseae significantly alleviated tomato (Solanum lycopersicum Mill.) early blight disease caused by Alternaria solani Sorauer. AMF pre-inoculation led to significant increases in activities of β-1,3-glucanase, chitinase, phenylalanine ammonia-lyase (PAL) and lipoxygenase (LOX) in tomato leaves upon pathogen inoculation. Mycorrhizal inoculation alone did not influence the transcripts of most genes tested. However, pathogen attack on AMF-inoculated plants provoked strong defense responses of three genes encoding pathogenesis-related proteins, PR1, PR2, and PR3, as well as defense-related genes LOX, AOC, and PAL, in tomato leaves. The induction of defense responses in AMF pre-inoculated plants was much higher and more rapid than that in un-inoculated plants in present of pathogen infection. Three tomato genotypes: a Castlemart wild-type (WT) plant, a jasmonate (JA) biosynthesis mutant (spr2), and a prosystemin-overexpressing 35S::PS plant were used to examine the role of the JA signaling pathway in AMF-primed disease defense. Pathogen infection on mycorrhizal 35S::PS plants led to higher induction of defense-related genes and enzymes relative to WT plants. However, pathogen infection did not induce these genes and enzymes in mycorrhizal spr2 mutant plants. Bioassays showed that 35S::PS plants were more resistant and spr2 plants were more susceptible to early blight compared with WT plants. Our finding indicates that mycorrhizal colonization enhances tomato resistance to early blight by priming systemic defense response, and the JA signaling pathway is essential for mycorrhiza-primed disease resistance.
Project description:Arbuscular mycorrhizal symbiosis is a mutualistic interaction between most land plants and fungi of the glomeromycotina subphylum. The initiation, development and regulation of this symbiosis involve numerous signalling events between and within the symbiotic partners. Among other signals, phytohormones are known to play important roles at various stages of the interaction. During presymbiotic steps, plant roots exude strigolactones which stimulate fungal spore germination and hyphal branching, and promote the initiation of symbiosis. At later stages, different plant hormone classes can act as positive or negative regulators of the interaction. Although the fungus is known to reciprocally emit regulatory signals, its potential contribution to the phytohormonal pool has received little attention, and has so far only been addressed by indirect assays. In this study, using mass spectrometry, we analyzed phytohormones released into the medium by germinated spores of the arbuscular mycorrhizal fungus Rhizophagus irregularis. We detected the presence of a cytokinin (isopentenyl adenosine) and an auxin (indole-acetic acid). In addition, we identified a gibberellin (gibberellin A4) in spore extracts. We also used gas chromatography to show that R. irregularis produces ethylene from methionine and the α-keto γ-methylthio butyric acid pathway. These results highlight the possibility for AM fungi to use phytohormones to interact with their host plants, or to regulate their own development.
Project description:Tea has been gaining increasing popularity all over the world in recent years, and its yield and quality depend on the growth and development of tea plants [Camellia sinensis (L.) Kuntze] in various environments. Nowadays, biotic stress and extreme weather, such as high temperature, drought, waterlogging, pests, and diseases, bring about much pressure on the production of tea with high quality. Wherein anthracnose, which is the most common and serious disease of tea plants, has earned more and more attention, as its control mainly relies on chemical pesticides. Arbuscular mycorrhizal fungi (AMF), forming symbiosis with most terrestrial plants, participate in plant resistance against the anthracnose disease, which was found by previous studies in a few herbaceous plants. However, there are a few studies about arbuscular mycorrhizal (AM) fungal regulation of the resistance to the anthracnose pathogen in woody plants so far. In this paper, we investigated the effect of AMF on the development of anthracnose caused by Colletotrichum camelliae and tried to decipher the pertinent mechanism through transcriptome analysis. Results showed that inoculating AMF significantly reduced the damage of anthracnose on tea seedlings by reducing the lesion area by 35.29% compared to that of the control. The content of superoxide anion and activities of catalase and peroxidase significantly increased (P < 0.05) in mycorrhizal treatment in response to the pathogen with 1.23, 2.00, and 1.39 times higher, respectively, than those in the control. Pathways of plant hormone signal transduction, mitogen-activated protein kinase (MAPK) signaling, and phenylpropanoid biosynthesis might play roles in this regulation according to the transcriptomic results. Further redundancy analysis (RDA) and partial least squares structural equation modeling (PLS-SEM) analysis found that plant hormones, such as auxin and ethylene, and the antioxidant system (especially peroxidase) were of great importance in the AM fungal alleviation of anthracnose. Our results preliminarily indicated the mechanisms of enhanced resistance in mycorrhizal tea seedlings to the anthracnose pathogen and provided a theoretical foundation for the application of AMF as one of the biological control methods in tea plantations.
Project description:The virtual transcriptome of G. intraradices (based on >430,000 reads) served as the basis for the design of an EST expresson array and for a number of analyses of gene expression in germinating spores, extraradical mycelium (ERM) and symbiotic root tissues. The G. intraradices EST expression array (4 x 72K) manufactured by Roche NimbleGen Systems Limited (Madison, WI) (http://www.nimblegen.com/products/exp/index.html) contained three independent, non identical, 60-mer probes per sequence. Included in the oligoarray were 22,402 G. intraradices sequences, 5785 random 60-mer control probes and labeling controls. We performed 12 hybridisations with samples from germinating spores (three biological replicates), extraradical mycelium (three biological replicates), symbiotic root tissues from Medicago (three biological replicates) and rice (1 replicate) as well as from microdissected arbuscule-colonized cortical cells of Medicago and rice (1 replicate each).