Arbuscular Mycorrhiza Development Research Articles

Long-lasting impact of chito-oligosaccharide application on strigolactone biosynthesis and fungal accommodation promotes arbuscular mycorrhiza in Medicago truncatula.

• The establishment of arbuscular mycorrhiza (AM) between plants and Glomeromycotina fungi is preceded by the exchange of chemical signals: fungal released Myc-factors, including chitoligosaccharides (CO) and lipo-chitooligosaccharides (LCO), activate plant symbiotic responses, while root exuded strigolactones stimulate hyphal branching and boost CO release. Furthermore, fungal signaling reinforcement through CO application was shown to promote AM development in Medicago truncatula, but the cellular and molecular bases of this effect remained unclear. • Here we focused on long-term M. truncatula responses to CO treatment, demonstrating its impact on the transcriptome of both mycorrhizal and non-mycorrhizal roots over several weeks and providing an insight into the mechanistic bases of the CO-dependent promotion of AM colonization. • CO treatment caused the long-lasting regulation of strigolactone biosynthesis and fungal accommodation related genes. This was mirrored by an increase in root didehydro-orobanchol content, and the promotion of accommodation responses to AM fungi in root epidermal cells. Lastly, an advanced down-regulation of AM symbiosis marker genes was observed at the latest time point in CO-treated plants, in line with an increased number of senescent arbuscules. • Overall, CO treatment triggered molecular, metabolic and cellular responses underpinning a protracted acceleration of AM development.

The Role of Medicago lupulina Interaction with Rhizophagus irregularis in the Determination of Root Metabolome at Early Stages of AM Symbiosis.

The nature of plant–fungi interaction at early stages of arbuscular mycorrhiza (AM) development is still a puzzling problem. To investigate the processes behind this interaction, we used the Medicago lupulina MlS-1 line that forms high-efficient AM symbiosis with Rhizophagus irregularis. AM fungus actively colonizes the root system of the host plant and contributes to the formation of effective AM as characterized by a high mycorrhizal growth response (MGR) in the host plant. The present study is aimed at distinguishing the alterations in the M. lupulina root metabolic profile as an indicative marker of effective symbiosis. We examined the root metabolome at the 14th and 24th day after sowing and inoculation (DAS) with low substrate phosphorus levels. A GS-MS analysis detected 316 metabolites. Results indicated that profiles of M. lupulina root metabolites differed from those in leaves previously detected. The roots contained fewer sugars and organic acids. Hence, compounds supporting the growth of mycorrhizal fungus (especially amino acids, specific lipids, and carbohydrates) accumulated, and their presence coincided with intensive development of AM structures. Mycorrhization determined the root metabolite profile to a greater extent than host plant development. The obtained data highlight the importance of active plant–fungi metabolic interaction at early stages of host plant development for the determination of symbiotic efficiency.

Multifarious and Interactive Roles of GRAS Transcription Factors During Arbuscular Mycorrhiza Development

Arbuscular mycorrhiza (AM) is a mutualistic symbiotic interaction between plant roots and AM fungi (AMF). This interaction is highly beneficial for plant growth, development and fitness, which has made AM symbiosis the focus of basic and applied research aimed at increasing plant productivity through sustainable agricultural practices. The creation of AM requires host root cells to undergo significant structural and functional modifications. Numerous studies of mycorrhizal plants have shown that extensive transcriptional changes are induced in the host during all stages of colonization. Advances have recently been made in identifying several plant transcription factors (TFs) that play a pivotal role in the transcriptional regulation of AM development, particularly those belonging to the GRAS TF family. There is now sufficient experimental evidence to suggest that GRAS TFs are capable to establish intra and interspecific interactions, forming a transcriptional regulatory complex that controls essential processes in the AM symbiosis. In this minireview, we discuss the integrative role of GRAS TFs in the regulation of the complex genetic re-programming determining AM symbiotic interactions. Particularly, research being done shows the relevance of GRAS TFs in the morphological and developmental changes required for the formation and turnover of arbuscules, the fungal structures where the bidirectional nutrient translocation occurs.

Sugar transporters of the SWEET family and their role in arbuscular mycorrhiza.

Plant sugar transporters play an essential role in the organism’s productivity by carrying out carbohydrate transportation from source cells in the leaves to sink cells in the cortex. In addition, they aid in the regulation of a substantial part of the exchange of nutrients with microorganisms in the rhizosphere (bacteria and fungi), an ty essential to the formation of symbiotic relationships. This review pays special attention to carbohydrate nutrition during the development of arbuscular mycorrhiza (AM), a symbiosis of plants with fungi from the Glomeromycotina subdivision. This relationship results in the host plant receiving micronutrients from the mycosymbiont, mainly phosphorus, and the fungus receiving carbon assimilation products in return. While the eff icient nutrient transport pathways in AM symbiosis are yet to be discovered, SWEET sugar transporters are one of the three key families of plant carbohydrate transporters. Specif ic AM symbiosis transporters can be identif ied among the SWEET proteins. The survey provides data on the study history, structure and localization, phylogeny and functions of the SWEET proteins. A high variability of both the SWEET proteins themselves and their functions is noted along with the fact that the same proteins may perform different functions in different plants. A special role is given to the SWEET transporters in AM development. SWEET transporters can also play a key role in abiotic stress tolerance, thus allowing plants to adapt to adverse environmental conditions. The development of knowledge about symbiotic systems will contribute to the creation of microbial preparations for use in agriculture in the Russian Federation.

Andrea Genre.

Andrea Genre.

Metabolic Alterations in Pisum sativum Roots during Plant Growth and Arbuscular Mycorrhiza Development.

Intensive exchange of nutrients is a crucial part of the complex interaction between a host plant and fungi within arbuscular mycorrhizal (AM) symbiosis. For the first time, the present study demonstrates how inoculation with AMF Rhizophagus irregularis affects the pea (Pisum sativum L.) root metabolism at key stages of plant development. These correspond to days 21 (vegetation), 42 (flowering initiation), and 56 (fruiting-green pod). Metabolome profiling was carried out by means of a state-of-the-art GC-MS technique. The content shifts revealed include lipophilic compounds, sugars, carboxylates, and amino acids. The metabolic alterations were principally dependent on the stage of plant development but were also affected by the development of AM fungi, a fact which highlights interaction between symbiotic partners. The comparison of the present data with the results of leaf metabolome profiling earlier obtained did not reveal common signatures of metabolic response to mycorrhization in leaves and roots. We supposed that the feedback for the development and symbiotic interaction on the part of the supraorganismic system (root + AM fungi) was the cause of the difference between the metabolic profile shift in leaf and root cells that our examination revealed. New investigations are required to expand our knowledge of metabolome plasticity of the whole organism and/or system of organisms, and such results might be put to use for the intensification of sustainable agriculture.

ОСОБЛИВОСТІ АДАПТИВНИХ СТРАТЕГІЙ ВИДІВ РОДУ GENTIANA L. В УМОВАХ ВИСОКОГІР’Я УКРАЇНСЬКИХ КАРПАТ

Peculiarities of adaptive strategies of rare species of Gentiana lutea L., Gentiana punctata L., Gentiana acaulis L., which allow them to survive in the highlands of the Ukrainian Carpathians, are analyzed. The ability of these species to survive in the highlands is determined not by their individual adaptive properties, but by the integrated interaction of morphological, anatomical, physiological, biorhythmological, reproductive and other features, which increase the adaptability of plants to specific habitats and shape their individual survival strategies.
 The findings show that the formation of semi-rosette structures in plant species of G. lutea, G. punctata, and G. acaulis is a morphological property that allows them to exist in specific conditions of the highlands, where the surface layer of air is heated better. A number of passive and active mechanisms of protection of generative organs are revealed.
 The upper leaves of monocarpic shoots of G. lutea and G. punctata together with the membranous cups create a kind of "micro-greenhouse" that protects the ovary from temperature changes. Both species are characterized by alternation of flowers in the inflorescence, which reduces the risk of damage in case of sudden frosts and belongs to the group of active adaptations that provide a favorable temperature for the development of generative organs of plants.
 The corolla of G. punctata plants is densely covered with dark spots, and in G. acaulis plants it has a bluish color with olive-green spots. This is a sign of high content of anthocyanins, the biosynthesis of which is enhanced at low temperatures and increase the background of UV radiation and partially compensates for the lack of thermal energy in plants and protects them from short-wave radiation.
 There is a correlation between the size of the corolla in plants and the hypsometric level of growth of species: increase in altitude above the sea level leads to size increase too. All species are characterized by a break in flowering and high life expectancy, which allows plants to restore the supply of nutrients necessary to ensure the processes of life, including seed production during a short-term vegetation period. These species are also characterized by miniaturization of morphometric parameters, which is a manifestation not only of interspecific features, but is also typical of the given species due to the localization of its populations at different hypsometric levels. The study demonstrated that differences in living conditions also affected the duration of functioning of rosette leaves. In species with lower hypsometric growth levels (G. lutea and G. punctata) only immature plants belong to the group of evergreen plants, and in the alpine species of G. acaulis - plants of all stages of ontogenesis. This allows plants to carry out photosynthesis immediately after the snow melts. The research findings reveal that the growing conditions also affected the morphology and anatomy of underground organs, which in G. lutea and G. punctata are represented by perennial additional storage roots with strong parenchymal secondary wood and interxylary phloem. In G. acaulis, the root system is represented by thin additional roots, which are characterized by moderate parenchymatization of the secondary phloem and xylem. Accordingly, it affects the storage of nutrients, the depot of which in G. lutea and G. punctata has a root system, and in G. acaulis - a rosette of leaves. The peculiarity of these species is the absence of root hairs, which is compensated by the development of arbuscular mycorrhiza, which is the result of symbiosis with glomus fungi (Glomeromycota).

AM-Induced Alteration in the Expression of Genes, Encoding Phosphorus Transporters and Enzymes of Carbohydrate Metabolism in Medicago lupulina.

Plant–microbe interactions, including those of arbuscular mycorrhiza (AM), have been investigated for a wide spectrum of model plants. The present study focuses on an analysis of gene expression that encodes phosphate and sugar transporters and carbohydrate metabolic enzymes in a new model plant, the highly mycotrophic Medicago lupulina MLS-1 line under conditions of phosphorus deficiency and inoculation with Rhizophagus irregularis. Expression profiles were detected by RT-PCR at six plant stages of development (second leaf, third leaf, shooting, axillary shoot branching initiation, axillary shoot branching, flowering initiation). In comparison to control (without AM), the variant with AM inoculation exhibited a significant elevation of transcription levels of carbohydrate metabolic enzymes (MlSUS, MlHXK1) and sucrose transporters (MlSUC4) in M. lupulina leaves at the shooting stage. We suggest that this leads to a significant increase in the frequency of AM infection, an abundance of mycelium in roots and an increase in AM efficiency (which is calculated by the fresh weight of aerial parts and roots at the axillary shoot branching initiation stage). In roots, the specificity of MlPT4 and MlATP1 gene expressions were revealed for effective AM symbiosis. The level of MlPT4 transcripts in AM roots increased more than tenfold in comparison to that of non-specific MlPT1 and MlPT2. For the first time, MlPT1 expression was shown to increase sharply against MlPT2 in M. lupulina roots without AM at the shooting initiation stage. A significant increase in MlRUB expression was revealed at late stages in the host plant’s development, during axillary shoot branching and flowering initiation. The opposite changes characterized MlHXK1 expression. Alteration in MlHXK1 gene transcription was the same, but was more pronounced in roots. The obtained results indicate the importance of genes that encode phosphate transporters and the enzymes of carbohydrate metabolism for effective AM development at the shooting stage in the host plant.

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.

Metabolic alterations in pea leaves during arbuscular mycorrhiza development.

Arbuscular mycorrhiza (AM) is known to be a mutually beneficial plant-fungal symbiosis; however, the effect of mycorrhization is heavily dependent on multiple biotic and abiotic factors. Therefore, for the proper employment of such plant-fungal symbiotic systems in agriculture, a detailed understanding of the molecular basis of the plant developmental response to mycorrhization is needed. The aim of this work was to uncover the physiological and metabolic alterations in pea (Pisum sativum L.) leaves associated with mycorrhization at key plant developmental stages. Plants of pea cv. Finale were grown in constant environmental conditions under phosphate deficiency. The plants were analyzed at six distinct time points, which corresponded to certain developmental stages of the pea: I: 7 days post inoculation (DPI) when the second leaf is fully unfolded with one pair of leaflets and a simple tendril; II: 21 DPI at first leaf with two pairs of leaflets and a complex tendril; III: 32 DPI when the floral bud is enclosed; IV: 42 DPI at the first open flower; V: 56 DPI when the pod is filled with green seeds; and VI: 90–110 DPI at the dry harvest stage. Inoculation with Rhizophagus irregularis had no effect on the fresh or dry shoot weight, the leaf photochemical activity, accumulation of chlorophyll a, b or carotenoids. However, at stage III (corresponding to the most active phase of mycorrhiza development), the number of internodes between cotyledons and the youngest completely developed leaf was lower in the inoculated plants than in those without inoculation. Moreover, inoculation extended the vegetation period of the host plants, and resulted in increase of the average dry weight per seed at stage VI. The leaf metabolome, as analyzed with GC-MS, included about three hundred distinct metabolites and showed a strong correlation with plant age, and, to a lesser extent, was influenced by mycorrhization. Metabolic shifts influenced the levels of sugars, amino acids and other intermediates of nitrogen and phosphorus metabolism. The use of unsupervised dimension reduction methods showed that (i) at stage II, the metabolite spectra of inoculated plants were similar to those of the control, and (ii) at stages IV and V, the leaf metabolic profiles of inoculated plants shifted towards the profiles of the control plants at earlier developmental stages. At stage IV the inoculated plants exhibited a higher level of metabolism of nitrogen, organic acids, and lipophilic compounds in comparison to control plants. Thus, mycorrhization led to the retardation of plant development, which was also associated with higher seed biomass accumulation in plants with an extended vegetation period. The symbiotic crosstalk between host plant and AM fungi leads to alterations in several biochemical pathways the details of which need to be elucidated in further studies.

A set of Arabidopsis genes involved in the accommodation of the downy mildew pathogen Hyaloperonospora arabidopsidis.

The intracellular accommodation structures formed by plant cells to host arbuscular mycorrhiza fungi and biotrophic hyphal pathogens are cytologically similar. Therefore we investigated whether these interactions build on an overlapping genetic framework. In legumes, the malectin-like domain leucine-rich repeat receptor kinase SYMRK, the cation channel POLLUX and members of the nuclear pore NUP107-160 subcomplex are essential for symbiotic signal transduction and arbuscular mycorrhiza development. We identified members of these three groups in Arabidopsis thaliana and explored their impact on the interaction with the oomycete downy mildew pathogen Hyaloperonospora arabidopsidis (Hpa). We report that mutations in the corresponding genes reduced the reproductive success of Hpa as determined by sporangiophore and spore counts. We discovered that a developmental transition of haustorial shape occurred significantly earlier and at higher frequency in the mutants. Analysis of the multiplication of extracellular bacterial pathogens, Hpa-induced cell death or callose accumulation, as well as Hpa- or flg22-induced defence marker gene expression, did not reveal any traces of constitutive or exacerbated defence responses. These findings point towards an overlap between the plant genetic toolboxes involved in the interaction with biotrophic intracellular hyphal symbionts and pathogens in terms of the gene families involved.

Kalliope Papadopoulou.

Kalliope Papadopoulou.

Medicago lupulina lines with defects in the development of efficient arbuscular mycorrhiza

Background. The work is aimed to solve actual problems in biology of arbuscular mycorrhiza (AM). Currently, a lot of mutants had been obtained in various plant model objects with defects in genes controlling AM development, however, the mechanisms controlling development of effective AM symbiosis are still unclear.
 Materials and methods. The authors conducted a mutagenesis in Medicago lupulina, a new convenient model plant for molecular-genetic studies. High mycotrophic M. lupulina line have early and high response to mycorrhization, high seed production, as well as signs of dwarfism under conditions without of AM and low level of phosphorus available for plants. This method allows visually to identify plant lines with defects in AM symbiosis.
 Results. 14 modes for mutagenesis by ethylmethanesulfonate were conducted. Usage of 3 mutagenesis modes allowed to obtain: productive M1 progeny with high part of viable seedlings (73.3%86.0%); 1405 plants in M2 progeny.
 Conclusion. According to population analysis for mutant plants in M2 progeny (up to M9 generation) 15 plant lines were selected: one Myc plant line unable to form AM symbiosis, 4 Pen plant lines unable to form AM symbiosis, but characterized by appressoria formation; 3 Rmd plant lines forming low-activity ineffective AM symbiosis; 3 Rmd plant lines forming low-activity effective AM and 4 Rmd++ plant lines forming effective AM with high abundance of symbiotic structures (mycelium/arbuscules/vesicles) in the roots.

Strigolactones as Regulators of Symbiotrophy of Plants and Microorganisms

Strigolactones (SLs) are signal molecules of a butenolide nature from the apocarotenoids family that are synthesized by plant organisms. They are widely propagated in nature and control different aspects of plant development, which allows us to consider them as phytohormones. It is known that SLs production and release into the rhizosphere are enhanced under a deficit of basic elements of plant nutrition. In addition, these compounds carry out the signaling function in the course of establishing supraorganismal relationships upon parasitism and symbiosis. In this review, the role of SLs in the formation of arbuscular mycorrhiza (AM), which provides a symbiotic pathway of Pi uptake by plants and partly nitrogen-fixing legume nodules, is examined. The data on SLs impact on AM fungi (AMFs) development at presymbiotic and symbiotic stages of plant growth were generalized. The main peculiarities of SLs structure providing their efficiency as “branching factors” of AMF hyphae and basic mechanisms of the possible influence of SLs on AMFs, including stimulation of mitochondria biogenesis, were described. Analysis of these data and phenotypes of plant mutants with disturbances in SLs biosynthesis and its regulation and also SLs perception and transport leads to the conclusion that their role in AM development is predominantly revealed in induction of AMF hyphae branching at the presymbiotic plant growth stage and is associated with plant response to Pi deficit. Analysis of the role these components of common symbiotic signaling cascade play in the regulation of SLs biosynthesis in the course of AM development and nitrogen-fixing legumes nodules was carried out. An important role of SLs in nodule development likely caused by their endogenous influence on its organogenesis was demonstrated. A possibility of the presence of different pathways usable by a plant upon AM and nodule development for activation of common symbiotic transcriptional factors, NSP1 and NSP2, participating in the regulation of SLs biosynthesis is discussed. The data concerning structural specificity of SLs and the results of phylogenetic analysis of genes encoding different components of SLs biosynthesis and signaling pathways and also symbiotic signaling cascade in plants provide evidence for the possible transformation of SLs signaling function from the hormonal one manifested inside the plant into the communicative one providing the establishment of interorganismal relations.

Transcriptional Regulation of Arbuscular Mycorrhiza Development.

Arbuscular mycorrhiza (AM) is an ancient symbiosis between land plants and fungi of the glomeromycotina that is widespread in the plant kingdom. AM improves plant nutrition, stress resistance and general plant performance, and thus represents a promising addition to sustainable agricultural practices. In return for delivering mineral nutrients, the obligate biotrophic AM fungi receive up to 20% of the photosynthetically fixed carbon from the plant. AM fungi colonize the inside of roots and form highly branched tree-shaped structures, called arbuscules, in cortex cells. The pair of the arbuscule and its host cell is considered the central functional unit of the symbiosis as it mediates the bidirectional nutrient exchange between the symbionts. The development and spread of AM fungi within the root is predominantly under the control of the host plant and depends on its developmental and physiological status. Intracellular accommodation of fungal structures is enabled by the remarkable plasticity of plant cells, which undergo drastic subcellular rearrangements. These are promoted and accompanied by cell-autonomous transcriptional reprogramming. AM development can be dissected into distinct stages using plant mutants. Progress in the application of laser dissection technology has allowed the assignment of transcriptional responses to specific stages and cell types. The first transcription factors controlling AM-specific gene expression and AM development have been discovered, and cis-elements required for AM-responsive promoter activity have been identified. An understanding of their connectivity and elucidation of transcriptional networks orchestrating AM development can be expected in the near future.

Computational investigation of small RNAs in the establishment of root nodules and arbuscular mycorrhiza in leguminous plants.

Many small RNAs have been confirmed to play important roles in the development of root nodules and arbuscular mycorrhiza. In this study, we carried out the identification of certain small RNAs in leguminous plants (Medicago truncatula, soybean, peanut and common bean), such as miRNAs, tRFs and srRNAs, as well as the computational investigation of their regulations. Thirty miRNAs were predicted to be involved in establishing root nodules and mycorrhiza, and 12 of them were novel in common bean and peanut. The generation of tRFs in M. truncatula was not associated with tRNA gene frequencies and codon usage. Six tRFs exhibited different expressions in mycorrhiza and root nodules. Moreover, srRNA5.8S in M. truncatula was generated from the regions with relatively low conservation at the rRNA 3' terminal. The protein-protein interactions between the proteins encoded by the target genes of miRNAs, tRFs and srRNAs were computed. The regulation of these three types of sRNAs in the symbiosis between leguminous plants and microorganisms is not a single regulation of certain signaling or metabolic pathways but a global regulation for the plants to own growth or specific events in symbiosis.

Plant growth and arbuscular mycorrhizae development in oil sands processing by-products

Soil pollutants such as hydrocarbons can induce toxic effects in plants and associated arbuscular mycorrhizal fungi (AMF). This study was conducted to evaluate if the legume Lotus corniculatus and the grass Elymus trachycaulus and arbuscular mycorrhizal fungi could grow in two oil sands processing by-products after bitumen extraction from the oil sands in northern Alberta, Canada. Substrate treatments were coarse tailings sand (CTS), a mix of dry mature fine tailings (MFT) with CTS (1:1) and Pleistocene sandy soil (hydrocarbon free); microbial treatments were without AMF, with AMF and AMF plus soil bacteria isolated from oil sands reclamation sites. Plant biomass, root morphology, leaf water content, shoot tissue phosphorus content and mycorrhizal colonization were evaluated. Both plant species had reduced growth in CTS and tailings mix relative to sandy soil. AMF frequency and intensity in roots of E. trachycaulus was not influenced by soil hydrocarbons; however, it decreased significantly over time in roots of L. corniculatus without bacteria in CTS. Mycorrhizal inoculation alone did not significantly improve plant growth in CTS and tailings mix; however, inoculation with mycorrhizae plus bacteria led to a significantly positive response of both plant species in CTS. Thus, combined inoculation with selected mycorrhizae and bacteria led to synergistic effects. Such combinations may be used in future to improve plant growth in reclamation of CTS and tailings mix.

The effect of inoculation with arbuscular mycorrhizal fungus Rhizophagus irregularis on cytokinin content in a highly mycotrophic Medicago lupulina line under low phosphorus level in the soil

The study is focused on the elucidation of the role of cytokinins (CKs, zeatin and zeatin riboside) in the development of effective arbuscular mycorrhiza (AM) symbiosis with Medicago lupulina. An important mechanism involved in the regulation of host plant growth is supposed to be linked to the modulation of plant hormone balance. The data obtained revealed the formation of an effective AM-symbiosis (M. lupulina + Rhizophagus irregularis) under phosphorus-deficiency. At the shooting stage (35<sup>th</sup> day after sowing), it is characterized by a decrease in the root:shoot ratio, the lowering in arbuscules and vesicle abundances, but an increase in the intensity of mycelium development. Mycorrhized plants differed from the control ones by higher CK levels in both roots and leaves. Zeatin and zeatin riboside concentration exhibited uneven alterations over time. A role of mycelium in the modulation of CK balance has been discussed.

Gene expression analyses in tomato near isogenic lines provide evidence for ethylene and abscisic acid biosynthesis fine-tuning during arbuscular mycorrhiza development.

Plant responses to the environment and microorganisms, including arbuscular mycorrhizal fungi, involve complex hormonal interactions. It is known that abscisic acid (ABA) and ethylene may be involved in the regulation of arbuscular mycorrhiza (AM) and that part of the detrimental effects of ABA deficiency in plants is due to ethylene overproduction. In this study, we aimed to determine whether the low susceptibility to mycorrhizal colonization in ABA-deficient mutants is due to high levels of ethylene and whether AM development is associated with changes in the steady-state levels of transcripts of genes involved in the biosynthesis of ethylene and ABA. For that, tomato (Solanum lycopersicum) ethylene overproducer epinastic (epi) mutant and the ABA-deficient notabilis (not) and sitiens (sit) mutants, in the same Micro-Tom (MT) genetic background, were inoculated with Rhizophagus clarus, and treated with the ethylene biosynthesis inhibitor aminoethoxyvinylglycine (AVG). The development of AM, as well as the steady-state levels of transcripts involved in ethylene (LeACS2, LeACO1 and LeACO4) and ABA (LeNCED) biosynthesis, was determined. The intraradical colonization in epi, not and sit mutants was significantly reduced compared to MT. The epi mutant completely restored the mycorrhizal colonization to the levels of MT with the application of 10µM of AVG, probably due to the inhibition of the ACC synthase gene expression. The steady-state levels of LeACS2 and LeACO4 transcripts were induced in mycorrhizal roots of MT, whereas the steady-state levels of LeACO1 and LeACO4 transcripts were significantly induced in sit, and the steady-state levels of LeNCED transcripts were significantly induced in all genotypes and in mycorrhizal roots of epi mutants treated with AVG. The reduced mycorrhizal colonization in sit mutants seems not to be limited by ethylene production via ACC oxidase regulation. Both ethylene overproduction and ABA deficiency impaired AM fungal colonization in tomato roots, indicating that, besides hormonal interactions, a fine-tuning of each hormone level is required for AM development.

Transcriptional profiling of arbuscular mycorrhizal roots exposed to high levels of phosphate reveals the repression of cell cycle-related genes and secreted protein genes in Rhizophagus irregularis.

The development of arbuscular mycorrhiza (AM) is strongly suppressed under high-phosphate (Pi) conditions. To investigate AM fungal responses during the suppression of AM by high Pi, we performed an RNA-seq analysis of Rhizophagus irregularis colonizing Lotus japonicus roots at different levels of Pi (20, 100, 300, and 500μM). AM fungal colonization decreased markedly under high-Pi conditions. In total, 163 fungal genes were differentially expressed among the four Pi treatments. Among these genes, a cell cycle-regulatory gene, cyclin-dependent kinase CDK1, and several DNA replication- and mitosis-related genes were repressed under high-Pi conditions. More than 20 genes encoding secreted proteins were also downregulated by high-Pi conditions, including the strigolactone-induced putative secreted protein 1 gene that enhances AM fungal colonization. In contrast, the expression of genes related to aerobic respiration and transport in R. irregularis were largely unaffected. Our data suggest that high Pi suppresses the expression of genes associated with fungal cell cycle progression or that encode secreted proteins that may be required for intercellular hyphal growth and arbuscule formation. However, high Pi has little effect on the transcriptional regulation of the primary metabolism or transport in preformed fungal structures.