Metabolic Profiling of Transgenic Tobacco Plants Synthesizing Bovine Interferon-Gamma

Interferon-gamma belongs to a large family of cytokines – multifunctional secreted proteins involved in animal non-specific immune response. Previously inbred lines of Nicotiana tabacum L. plants harboring a heterologous gene of bovine interferon-gamma Bt-sIFNG under the control of a constitutive 35S CaMV promoter have been created by Agrobacterium-mediated genetic transformation. The antiviral and immunomodulatory activities of plant-produced interferon-gamma in bovine cell culture and laboratory animals (mice) were observed. A state-of-the-art GS-MS technique has been used to identify the possible effect of the transformation on the plant’s metabolome. Total profiles included 350 metabolites from leaves, among which 150 substances were identified up to their class and 80 up to the exact metabolite. Metabolite profiling revealed that plants able to synthesize interferon-gamma are characterized by a higher level of amino acids and other substances involved in nitrogen metabolism. In transgenic plants intensification of the secondary metabolism was also detected. Some alterations were distinguished in plant metabolome depending on cultivation conditions

Evolution of 14-3-3 Proteins in Angiosperm Plants: Recurring Gene Duplication and Loss

14-3-3 proteins are key regulatory factors in plants and are involved in a broad range of physiological processes. We addressed the evolutionary history of 14-3-3s from 46 angiosperm species, including basal angiosperm Amborella and major lineage of monocotyledons and eudicotyledons. Orthologs of Arabidopsis isoforms were detected. There were several rounds of duplication events in the evolutionary history of the 14-3-3 protein family in plants. At least four subfamilies (iota, epsilon, kappa, and psi) formed as a result of ancient duplication in a common ancestor of angiosperm plants. Recent duplication events followed by gene loss in plant lineage, among others Brassicaceae, Fabaceae, and Poaceae, further shaped the high diversity of 14-3-3 isoforms in plants. Coexpression data showed that 14-3-3 proteins formed different functional groups in different species. In some species, evolutionarily related groups of 14-3-3 proteins had coexpressed together under certain physiological conditions, whereas in other species, closely related isoforms expressed in the opposite manner. A possible explanation is that gene duplication and loss is accompanied by functional plasticity of 14-3-3 proteins

Effects of Trophic Acclimation on Growth and Expression Profiles of Genes Encoding Enzymes of Primary Metabolism and Plastid Transporters of <i>Chlamydomonas reinhardtii</i>

In this paper, the effect of prolonged trophic acclimation on the subsequent growth of Chlamydomonas reinhardtii batch cultures was studied. The mixotrophic (light + acetate) acclimation stimulated subsequent growth at both mixotrophy and autotrophy conditions and altered the expression profile of genes encoding enzymes of primary metabolism and plastid transporters. Besides the trophic effect, the influence of Chlamydomonas culture growth stage on gene expression was determined. Under mixotrophic conditions, this effect was most pronounced in the first half of the exponential growth with partial retention of the previous acclimation period traits. The autotrophy acclimation effect was more complex and its significance was enhanced at the end of the growth and in the stationary phase

The Role of <i>Medicago lupulina</i> Interaction with <i>Rhizophagus irregularis</i> 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

Mycorrhiza-Induced Alterations in Metabolome of <i>Medicago lupulina</i> Leaves during Symbiosis Development

The present study is aimed at disclosing metabolic profile alterations in the leaves of the Medicago lupulina MlS-1 line that result from high-efficiency arbuscular mycorrhiza (AM) symbiosis formed with Rhizophagus irregularis under condition of a low phosphorus level in the substrate. A highly effective AM symbiosis was established in the period from the stooling to the shoot branching initiation stage (the efficiency in stem height exceeded 200%). Mycorrhization led to a more intensive accumulation of phosphates (glycerophosphoglycerol and inorganic phosphate) in M. lupulina leaves. Metabolic spectra were detected with GS-MS analysis. The application of complex mathematical analyses made it possible to identify the clustering of various groups of 320 metabolites and thus demonstrate the central importance of the carbohydrate and carboxylate-amino acid clusters. The results obtained indicate a delay in the metabolic development of mycorrhized plants. Thus, AM not only accelerates the transition between plant developmental stages but delays biochemical “maturation” mainly in the form of a lag of sugar accumulation in comparison with non-mycorrhized plants. Several methods of statistical modeling proved that, at least with respect to determining the metabolic status of host-plant leaves, stages of phenological development have priority over calendar age

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