Multiple control levels of root system remodeling in arbuscular mycorrhizal symbiosis.

In nature, the root systems of most plants develop intimate symbioses with glomeromycotan fungi that assist in the acquisition of mineral nutrients and water through uptake from the soil and direct delivery into the root cortex. Root systems are endowed with a strong, environment-responsive architectural plasticity that also manifests itself during the establishment of arbuscular mycorrhizal (AM) symbioses, predominantly in lateral root proliferation. In this review, we collect evidence for the idea that AM-induced root system remodeling is regulated at several levels: by AM fungal signaling molecules and by changes in plant nutrient status and distribution within the root system

Ramf: An Open-Source R Package for Statistical Analysis and Display of Quantitative Root Colonization by Arbuscular Mycorrhiza Fungi

Data analysis and graphical representation form an essential part of scientific research dissemination. The life-science community is moving towards a more transparent presentation of single data points or data distributions and away from mean values displayed as bar charts. To facilitate transparent data display to the mycorrhiza community, we present “Ramf” an open-source R package for statistical analysis and preparation of a variety of publication-ready plots, custom-made for analyzing and displaying quantitative root colonization by arbuscular mycorrhiza fungi or any kind of data to be displayed in the same format. Ramf replaces the scripting needed for data analysis and can be readily used by researchers not acquainted with R. In addition, the package is open to improvements by the community. Ramf is available at https://github.com/mchiapello/Ramf

A Flexible, Low-Cost Hydroponic Co-Cultivation System for Studying Arbuscular Mycorrhiza Symbiosis

Arbuscular mycorrhiza (AM) is a widespread symbiosis between plant roots and fungi of the Glomeromycotina, which improves nutrient uptake by plants. The molecular mechanisms underlying development and function of the symbiosis are subject to increasing research activity. Since AM occurs in the soil, most studies targeting a molecular understanding of AM development and function, use solid substrates for co-cultivating plants and AM fungi. However, for some experiments very clean roots, highly controlled nutrient conditions or applications of defined concentrations of signaling molecules (such as hormones) or other small chemicals (such as synthetic inhibitors or signaling agonists) are needed. To this end, hydroponics has been widely used in research on mechanisms of plant nutrition and some hydroponic systems were developed for AM fungal spore amplification. Here, we present a hydroponics set-up, which can be successfully utilized for experimental root colonization assays. We established a "tip-wick" system based on pipette tips and rock wool wicks for co-cultivation of AM fungi with small model plants such as Lotus japonicus. A larger "Falcon-wick" system using Falcon tubes and rockwool wicks was developed for larger model plants such as rice. The hydroponic system can also be employed for growing L. japonicus hairy roots after transformation by Agrobacterium rhizogenes, thus circumventing the laborious cultivation on agar medium-containing Petri dishes during hairy root development. The tip-wick and Falcon-wick systems are easy to use and can be built with low cost, conventional and reusable lab plastic ware and a simple aquarium pump

Tracking Lipid Transfer by Fatty Acid Isotopolog Profiling from Host Plants to Arbuscular Mycorrhiza Fungi

Lipid transfer from host plants to arbuscular mycorrhiza fungi was hypothesized for several years because sequenced arbuscular mycorrhiza fungal genomes lack genes encoding cytosolic fatty acid synthase (Wewer et al., 2014;Rich et al., 2017). It was finally shown by two independent experimental approaches (Jiang et al., 2017;Keymer et al., 2017;Luginbuehl et al., 2017). One approach used a technique called isotopolog profiling (Keymer et al., 2017). Isotopologs are molecules, which differ only in their isotopic composition. For isotopolog profiling an organism is fed with a heavy isotope labelled precursor metabolite. Subsequently, the labelled isotopolog composition of metabolic products is analysed via mass spectrometry. The detected isotopolog pattern of the metabolite(s) of interest yields information about metabolic pathways and fluxes (Ahmed et al., 2014). The following protocol describes an experimental setup, which enables separate isotopolog profiling of fatty acids in plant roots colonized by arbuscular mycorrhiza fungi and their associated fungal extraradical mycelium, to elucidate fluxes between both symbiotic organisms. We predict that this strategy can also be used to study metabolite fluxes between other organisms if the two interacting organisms can be physically separated

Full establishment of arbuscular mycorrhizal symbiosis in rice occurs independently of enzymatic jasmonate biosynthesis.

Development of the mutualistic arbuscular mycorrhiza (AM) symbiosis between most land plants and fungi of the Glomeromycota is regulated by phytohormones. The role of jasmonate (JA) in AM colonization has been investigated in the dicotyledons Medicago truncatula, tomato and Nicotiana attenuata and contradicting results have been obtained with respect to a neutral, promotive or inhibitory effect of JA on AM colonization. Furthermore, it is currently unknown whether JA plays a role in AM colonization of monocotyledonous roots. Therefore we examined whether JA biosynthesis is required for AM colonization of the monocot rice. To this end we employed the rice mutant constitutive photomorphogenesis 2 (cpm2), which is deficient in JA biosynthesis. Through a time course experiment the amount and morphology of fungal colonization did not differ between wild-type and cpm2 roots. Furthermore, no significant difference in the expression of AM marker genes was detected between wild type and cpm2. However, treatment of wild-type roots with 50 μM JA lead to a decrease of AM colonization and this was correlated with induction of the defense gene PR4. These results indicate that JA is not required for AM colonization of rice but high levels of JA in the roots suppress AM development likely through the induction of defense.CG was supported by a PhD fellowship of the German National Merit Foundation (Studienstiftung des Deutschen Volkes); http://www.studienstiftung.de/ and the Roche Foundation (Switzerland). Research in the UP laboratory was supported by the SNF 'professeur boursier' grants PP00A-110874 and PP00P3-130704; http://www.snf.ch/en/Pages/default.aspx. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.This is the final version of the article. It first appeared from PLOS via http://dx.doi.org/ 10.1371/journal.pone.012342

Divergence of Evolutionary Ways Among Common sym Genes: CASTOR and CCaMK Show Functional Conservation Between Two Symbiosis Systems and Constitute the Root of a Common Signaling Pathway

In recent years a number of legume genes involved in root nodule (RN) symbiosis have been identified in the model legumes, Lotus japonicus (Lotus) and Medicago truncatula. Among them, a distinct set of genes has been categorized as a common symbiosis pathway (CSP), because they are also essential for another mutual interaction, the arbuscular mycorrhiza (AM) symbiosis, which is evolutionarily older than the RN symbiosis and is widely distributed in the plant kingdom. Based on the concept that the legume RN symbiosis has evolved from the ancient AM symbiosis, one issue is whether the CSP is functionally conserved between non-nodulating plants, such as rice, and nodulating legumes. We identified three rice CSP gene orthologs, OsCASTOR, OsPOLLUX and OsCCaMK, and demonstrated the indispensable roles of OsPOLLUX and OsCCaMK in rice AM symbiosis. Interestingly, molecular transfection of either OsCASTOR or OsCCaMK could fully complement symbiosis defects in the corresponding Lotus mutant lines for both the AM and RN symbioses. Our results not only provide a conserved genetic basis for the AM symbiosis between rice and Lotus, but also indicate that the core of the CSP has been well conserved during the evolution of RN symbiosis. Through evolution, CASTOR and CCaMK have remained as the molecular basis for the maintenance of CSP functions in the two symbiosis system

Auxin Perception Is Required for Arbuscule Development in Arbuscular Mycorrhizal Symbiosis

Most land plant species live in symbiosis with arbuscular mycorrhizal fungi. These fungi differentiate essential functional structures called arbuscules in root cortical cells from which mineral nutrients are released to the plant. We investigated the role of microRNA393 (miR393), an miRNA that targets several auxin receptors, in arbuscular mycorrhizal root colonization. Expression of the precursors of the miR393 was down-regulated during mycorrhization in three different plant species: Solanum lycopersicum, Medicago truncatula, and Oryza sativa. Treatment of S. lycopersicum, M. truncatula, and O. sativa roots with concentrations of synthetic auxin analogs that did not affect root development stimulated mycorrhization, particularly arbuscule formation. DR5-GUS, a reporter for auxin response, was preferentially expressed in root cells containing arbuscules. Finally, overexpression of miR393 in root tissues resulted in down-regulation of auxin receptor genes (transport inhibitor response1 and auxin-related F box) and underdeveloped arbuscules in all three plant species. These results support the conclusion that miR393 is a negative regulator of arbuscule formation by hampering auxin perception in arbuscule-containing cells

Sculpting the soil microbiota

Soil is a living ecosystem, the health of which depends on fine interactions among its abiotic and biotic components. These form a delicate equilibrium maintained through a multilayer network that absorbs certain perturbations and guarantees soil functioning. Deciphering the principles governing the interactions within soils is of critical importance for their management and conservation. Here, we focus on soil microbiota and discuss the complexity of interactions that impact the composition and function of soil microbiota and their interaction with plants. We discuss how physical aspects of soils influence microbiota composition and how microbiota–plant interactions support plant growth and responses to nutrient deficiencies. We predict that understanding the principles determining the configuration and functioning of soil microbiota will contribute to the design of microbiota-based strategies to preserve natural resources and develop more environmentally friendly agricultural practices

Lotus japonicus karrikin receptors display divergent ligand-binding specificities and organ-dependent redundancy

Author summary Plant hormone signaling is crucial for development and for adequate responses to biotic and abiotic environmental conditions. The most recently discovered plant hormone receptor KARRIKIN INSENSITVE 2 (KAI2), binds a small butenolide called karrikin that was discovered in smoke and induces germination of fire-following plants. Several lines of evidence suggest a yet elusive endogenous hormone, which acts as ligand for KAI2. Until its identification, synthetic karrikins or the strigolactone-like molecule GR24 are used to probe the karrikin signaling pathway. While the model plant Arabidopsis contains only one KAI2 gene, several copies are maintained in other species suggesting sub-functionalization. We report that genomes of species in the legume hologalegina clade encode two KAI2 versions. In Lotus japonicus, they diverge in their binding ability to synthetic ligands due to three amino acid changes in their binding pocket, of which two are conserved across legumes and one has independently occurred in several species across the angiosperm phylogeny. Surprisingly, L. japonicus hypocotyls react with developmental responses to two different karrikins (KAR(1), KAR(2)) and a synthetic strigolactone rac-GR24, while root development responds only to KAR(1). This shows that there is not only diversity in ligand-receptor relationships but possibly also organ-specific uptake or metabolism of divergent butenolide molecules. Karrikins (KARs), smoke-derived butenolides, are perceived by the alpha/beta-fold hydrolase KARRIKIN INSENSITIVE2 (KAI2) and thought to mimic endogenous, yet elusive plant hormones tentatively called KAI2-ligands (KLs). The sensitivity to different karrikin types as well as the number of KAI2 paralogs varies among plant species, suggesting diversification and co-evolution of ligand-receptor relationships. We found that the genomes of legumes, comprising a number of important crops with protein-rich, nutritious seed, contain two or more KAI2 copies. We uncover sub-functionalization of the two KAI2 versions in the model legume Lotus japonicus and demonstrate differences in their ability to bind the synthetic ligand GR24(ent-5DS) in vitro and in genetic assays with Lotus japonicus and the heterologous Arabidopsis thaliana background. These differences can be explained by the exchange of a widely conserved phenylalanine in the binding pocket of KAI2a with a tryptophan in KAI2b, which arose independently in KAI2 proteins of several unrelated angiosperms. Furthermore, two polymorphic residues in the binding pocket are conserved across a number of legumes and may contribute to ligand binding preferences. The diversification of KAI2 binding pockets suggests the occurrence of several different KLs acting in non-fire following plants, or an escape from possible antagonistic exogenous molecules. Unexpectedly, L. japonicus responds to diverse synthetic KAI2-ligands in an organ-specific manner. Hypocotyl growth responds to KAR(1), KAR(2) and rac-GR24, while root system development responds only to KAR(1). This differential responsiveness cannot be explained by receptor-ligand preferences alone, because LjKAI2a is sufficient for karrikin responses in the hypocotyl, while LjKAI2a and LjKAI2b operate redundantly in roots. Instead, it likely reflects differences between plant organs in their ability to transport or metabolise the synthetic KLs. Our findings provide new insights into the evolution and diversity of butenolide ligand-receptor relationships, and open novel research avenues into their ecological significance and the mechanisms controlling developmental responses to divergent KLs