Performance of the Photosynthetic Apparatus under Glass with a Luminophore Modifying Red-To-Far-Red-Light Ratio-A Case Study.

The aim of this study was to examine the effect of the modified light spectrum of glass containing red luminophore on the performance of the photosynthetic apparatus of two types of lettuce cultivated in soil in a greenhouse. Butterhead and iceberg lettuce were cultivated in two types of greenhouses: (1) covered with transparent glass (control) and (2) covered with glass containing red luminophore (red). After 4 weeks of culture, structural and functional changes in the photosynthetic apparatus were examined. The presented study indicated that the red luminophore used changed the sunlight spectrum, providing an adequate blue:red light ratio, while decreasing the red:far-red radiation ratio. In such light conditions, changes in the efficiency parameters of the photosynthetic apparatus, modifications in the chloroplast ultrastructure, and altered proportions of structural proteins forming the photosynthetic apparatus were observed. These changes led to a decrease of CO2 carboxylation efficiency in both examined lettuce types.

The Adjustment Strategy of Venus Flytrap Photosynthetic Apparatus to UV-A Radiation.

The objective of this study was to investigate the response of the photosynthetic apparatus of the Venus flytrap (Dionaea muscipula J. Ellis) to UV-A radiation stress as well as the role of selected secondary metabolites in this process. Plants were subjected to 24 h UV-A treatment. Subsequently, chl a fluorescence and gas exchange were measured in living plants. On the collected material, analyses of the photosynthetic pigments and photosynthetic apparatus proteins content, as well as the contents and activity of selected antioxidants, were performed. Measurements and analyses were carried out immediately after the stress treatment (UV plants) and another 24 h after the termination of UV-A exposure (recovery plants). UV plants showed no changes in the structure and function of their photosynthetic apparatus and increased contents and activities of some antioxidants, which led to efficient CO2 carboxylation, while, in recovery plants, a disruption of electron flow was observed, resulting in lower photosynthesis efficiency. Our results revealed that D. muscipula plants underwent two phases of adjustment to UV-A radiation. The first was a regulatory phase related to the exploitation of available mechanisms to prevent the over-reduction of PSII RC. In addition, UV plants increased the accumulation of plumbagin as a potential component of a protective mechanism against the disruption of redox homeostasis. The second was an acclimatization phase initiated after the running down of the regulatory process and decrease in photosynthesis efficiency.

Response of physiological parameters in Dionaea muscipula J. Ellis teratomas transformed with rolB oncogene.

BACKGROUND: Plant transformation with rol oncogenes derived from wild strains of Rhizobium rhizogenes is a popular biotechnology tool. Transformation effects depend on the type of rol gene, expression level, and the number of gene copies incorporated into the plant's genomic DNA. Although rol oncogenes are known as inducers of plant secondary metabolism, little is known about the physiological response of plants subjected to transformation. RESULTS: In this study, the physiological consequences of rolB oncogene incorporation into the DNA of Dionaea muscipula J. Ellis was evaluated at the level of primary and secondary metabolism. Examination of the teratoma (transformed shoots) cultures of two different clones (K and L) showed two different strategies for dealing with the presence of the rolB gene. Clone K showed an increased ratio of free fatty acids to lipids, superoxide dismutase activity, synthesis of the oxidised form of glutathione, and total pool of glutathione and carotenoids, in comparison to non-transformed plants (control). Clone L was characterised by increased accumulation of malondialdehyde, proline, activity of superoxide dismutase and catalase, total pool of glutathione, ratio of reduced form of glutathione to oxidised form, and accumulation of selected phenolic acids. Moreover, clone L had an enhanced ratio of total triglycerides to lipids and accumulated saccharose, fructose, glucose, and tyrosine. CONCLUSIONS: This study showed that plant transformation with the rolB oncogene derived from R. rhizogenes induces a pleiotropic effect in plant tissue after transformation. Examination of D. muscipula plant in the context of transformation with wild strains of R. rhizogenes can be a new source of knowledge about primary and secondary metabolites in transgenic organisms.

Endopolyploidy Variation in Wild Barley Seeds across Environmental Gradients in Israel.

Wild barley is abundant, occupying large diversity of sites, ranging from the northern mesic Mediterranean meadows to the southern xeric deserts in Israel. This is also reflected in its wide phenotypic heterogeneity. We investigated the dynamics of DNA content changes in seed tissues in ten wild barley accessions that originated from an environmental gradient in Israel. The flow cytometric measurements were done from the time shortly after pollination up to the dry seeds. We show variation in mitotic cell cycle and endoreduplication dynamics in both diploid seed tissues (represented by seed maternal tissues and embryo) and in the triploid endosperm. We found that wild barley accessions collected at harsher xeric environmental conditions produce higher proportion of endoreduplicated nuclei in endosperm tissues. Also, a comparison of wild and cultivated barley strains revealed a higher endopolyploidy level in the endosperm of wild barley, that is accompanied by temporal changes in the timing of the major developmental phases. In summary, we present a new direction of research focusing on connecting spatiotemporal patterns of endoreduplication in barley seeds and possibly buffering for stress conditions.

Photosynthetic apparatus efficiency, phenolic acid profiling and pattern of chosen phytohormones in pseudometallophyte Alyssum montanum.

The present study investigated the response of non-metallicolous (NM) and metallicolous (M) Alyssum montanum shoots cultured in vitro on a medium supplemented simultaneously with heavy metals (HMs) to identify mechanisms involved in alleviating metal-induced damage. Plant status in respect to photosynthetic apparatus efficiency was determined and linked with changes in biochemical composition of shoots, namely phenolic acids' and stress-related phytohormones. Results showed the considerable inter-ecotype differences in (1) the photosynthetic pigments' amount, (2) the functioning of membrane electron transporters as well as (3) the linear and alternative electron transport pathways, whose lower values were reported in NM than in M HM-treated culture. Photosynthetic apparatus protection in M specimens was assured by the activation of cinnamic acid synthesis (by phenylalanine ammonia lyase) and its further transformations to benzoic acid derivatives with high ability to counteract oxidative stress, that was accompanied by the overexpression of jasmonic acid stimulating antioxidant machinery. In turn, detrimental HM effects on NM shoots could result from the diminution of most phenolics' accumulation, and only the content of coumarate (produced by bifunctional phenylalanine/tyrosine ammonia lyase) and rosmarinic acid increased. All these together with an enhanced concentration of abscisic acid might suggest that NM strategy to cope with HMs is based mostly on a restriction of metal movement with transpiration flow and their limited distribution in leaves. Summarizing, our findings for the first time point out the physiological and metabolic adaptation of pseudometallophyte A. montanum to adverse conditions.

Transformed tissue of Dionaea muscipula J. Ellis as a source of biologically active phenolic compounds with bactericidal properties.

The Venus flytrap (Dionaea muscipula J. Ellis) is a carnivorous plant able to synthesize large amounts of phenolic compounds, such as phenylpropanoids, flavonoids, phenolic acids, and 1,4-naphtoquinones. In this study, the first genetic transformation of D. muscipula tissues is presented. Two wild-type Rhizobium rhizogenes strains (LBA 9402 and ATCC 15834) were suitable vector organisms in the transformation process. Transformation led to the formation of teratoma (transformed shoot) cultures with the bacterial rolB gene incorporated into the plant genome in a single copy. Using high-pressure liquid chromatography, we demonstrated that transgenic plants were characterized by an increased quantity of phenolic compounds, including 1,4-naphtoquinone derivative, plumbagin (up to 106.63 mg × g-1 DW), and phenolic acids (including salicylic, caffeic, and ellagic acid), in comparison to non-transformed plants. Moreover, Rhizobium-mediated transformation highly increased the bactericidal properties of teratoma-derived extracts. The antibacterial properties of transformed plants were increased up to 33% against Staphylococcus aureus, Enterococcus faecalis, and Escherichia coli and up to 7% against Pseudomonas aeruginosa. For the first time, we prove the possibility of D. muscipula transformation. Moreover, we propose that transformation may be a valuable tool for enhancing secondary metabolite production in D. muscipula tissue and to increase bactericidal properties against human antibiotic-resistant bacteria. KEY POINTS: • Rhizobium-mediated transformation created Dionaea muscipula teratomas. • Transformed plants had highly increased synthesis of phenolic compounds. • The MBC value was connected with plumbagin and phenolic acid concentrations.

Stem Photosynthesis-A Key Element of Grass Pea (Lathyrus sativus L.) Acclimatisation to Salinity.

Grass pea (Lathyrus sativus) is a leguminous plant of outstanding tolerance to abiotic stress. The aim of the presented study was to describe the mechanism of grass pea (Lathyrus sativus L.) photosynthetic apparatus acclimatisation strategies to salinity stress. The seedlings were cultivated in a hydroponic system in media containing various concentrations of NaCl (0, 50, and 100 mM), imitating none, moderate, and severe salinity, respectively, for three weeks. In order to characterise the function and structure of the photosynthetic apparatus, Chl a fluorescence, gas exchange measurements, proteome analysis, and Fourier-transform infrared spectroscopy (FT-IR) analysis were done inter alia. Significant differences in the response of the leaf and stem photosynthetic apparatus to severe salt stress were observed. Leaves became the place of harmful ion (Na+) accumulation, and the efficiency of their carboxylation decreased sharply. In turn, in stems, the reconstruction of the photosynthetic apparatus (antenna and photosystem complexes) activated alternative electron transport pathways, leading to effective ATP synthesis, which is required for the efficient translocation of Na+ to leaves. These changes enabled efficient stem carboxylation and made them the main source of assimilates. The observed changes indicate the high plasticity of grass pea photosynthetic apparatus, providing an effective mechanism of tolerance to salinity stress.

Dynamics of endoreduplication in developing barley seeds.

Seeds are complex biological systems comprising three genetically distinct tissues: embryo, endosperm, and maternal tissues (including seed coats and pericarp) nested inside one another. Cereal grains represent a special type of seeds, with the largest part formed by the endosperm, a specialized triploid tissue ensuring embryo protection and nourishment. We investigated dynamic changes in DNA content in three of the major seed tissues from the time of pollination up to the dry seed. We show that the cell cycle is under strict developmental control in different seed compartments. After an initial wave of active cell division, cells switch to endocycle and most endoreduplication events are observed in the endosperm and seed maternal tissues. Using different barley cultivars, we show that there is natural variation in the kinetics of this process. During the terminal stages of seed development, specific and selective loss of endoreduplicated nuclei occurs in the endosperm. This is accompanied by reduced stability of the nuclear genome, progressive loss of cell viability, and finally programmed cell death. In summary, our study shows that endopolyploidization and cell death are linked phenomena that frame barley grain development.

Reactive oxygen species metabolism and photosynthetic performance in leaves of Hordeum vulgare plants co-infested with Heterodera filipjevi and Aceria tosichella.

KEY MESSAGE: Defence responses of cyst nematode and/or wheat curl mite infested barley engage the altered reactive oxygen species production, antioxidant machinery, carbon dioxide assimilation and photosynthesis efficiency. The primary aim of this study was to determine how barley responds to two pests infesting separately or at once; thus barley was inoculated with Heterodera filipjevi (Madzhidov) Stelter (cereal cyst nematode; CCN) and Aceria tosichella Keifer (wheat curl mite; WCM). To verify hypothesis about the involvement of redox metabolism and photosynthesis in barley defence responses, biochemical, photosynthesis efficiency and chlorophyll a fluorescence measurements as well as transmission electron microscopy were implemented. Inoculation with WCM (apart from or with CCN) brought about a significant suppression in the efficiency of electron transport outside photosystem II reaction centres. This limitation was an effect of diminished pool of rapidly reducing plastoquinone and decreased total electron carriers. Infestation with WCM (apart from or with CCN) also significantly restricted the electron transport on the photosystem I acceptor side, therefore produced reactive oxygen species oxidized lipids in cells of WCM and double infested plants and proteins in cells of WCM-infested plants. The level of hydrogen peroxide was significantly decreased in double infested plants because of glutathione-ascorbate cycle involvement. The inhibition of nitrosoglutathione reductase promoted the accumulation of S-nitrosoglutathione increasing antioxidant capacity in cells of double infested plants. Moreover, enhanced arginase activity in WCM-infested plants could stimulate synthesis of polyamines participating in plant antioxidant response. Infestation with WCM (apart from or with CCN) significantly reduced the efficiency of carbon dioxide assimilation by barley leaves, whereas infection only with CCN expanded photosynthesis efficiency. These were accompanied with the ultrastructural changes in chloroplasts during CCN and WCM infestation.

Elicitation-Based Method for Increasing the Production of Antioxidant and Bactericidal Phenolic Compounds in Dionaea muscipula J. Ellis Tissue.

The carnivorous plant Dionaea muscipula J. Ellis (Venus flytrap) is a widely known medical herb, capable of producing various phenolic compounds known for their strong antioxidant and antibacterial properties. In the pharmaceutical industry, Venus flytrap is grown in tissue cultures, as the natural population of D. muscipula is very limited. Here, we describe an improved method to increase the quantity and quality of phenolic compounds produced in D. muscipula. This is achieved by combining biotic elicitation (using Cronobacter sakazakii bacteria lysate) of D. muscipula cultured with rotary shaking (hydromechanical stress), which we describe here for the first time. The antibacterial activity and the antioxidant properties of the obtained compounds were studied on two antibiotic-resistant human pathogenic bacteria. The proposed plant culture conditions resulted in an increase in fresh weight, as well as a higher total phenolic content, in comparison to traditional tissue cultures on agar-solidified medium. With the use of high-performance liquid chromatography, we demonstrated that the described elicitation strategy leads to an increased synthesis of myricetin, caffeic acid, ellagic acid and plumbagin in D. muscipula tissue. We also found that a higher level of antioxidant activity, exhibited by the plant extract, corresponded with its higher phenylpropanoid content. The bactericidal activity of the extract against Staphylococcus aureus was dependent on the duration of plant culture under described elicitation conditions, whereas neither elicitation condition (duration or elicitor concentration) seemed relevant for the bactericidal activity of the extract towards Escherichia coli. This suggest that Gram-negative bacteria are less sensitive to compounds derived from Venus flytrap tissue.

Can Ceylon Leadwort (Plumbago zeylanica L.) Acclimate to Lead Toxicity?-Studies of Photosynthetic Apparatus Efficiency.

Ceylon leadwort (Plumbago zeylanica) is ornamental plant known for its pharmacological properties arising from the abundant production of various secondary metabolites. It often grows in lead polluted areas. The aim of presented study was to evaluate the survival strategy of P. zeylanica to lead toxicity via photosynthetic apparatus acclimatization. Shoots of P. zeylanica were cultivated on media with different Pb concentrations (0.0, 0.05, and 0.1 g Pb∙l-1). After a four-week culture, the efficiency of the photosynthetic apparatus of plants was evaluated by Chl a fluorescence measurement, photosynthetic pigment, and Lhcb1, PsbA, PsbO, and RuBisCo protein accumulation, antioxidant enzymes activity, and chloroplast ultrastructure observation. Plants from lower Pb concentration revealed no changes in photosynthetic pigments content and light-harvesting complex (LHCII) size, as well as no limitation on the donor side of Photosystem II Reaction Centre (PSII RC). However, the activity and content of antioxidant enzymes indicated a high risk of limitation on the acceptor side of Photosystem I. In turn, plants from 0.1 g Pb∙l-1 showed a significant decrease in pigments content, LHCII size, the amount of active PSII RC, oxygen-evolving complex activity, and significant remodeling of chloroplast ultrastructure indicated limitation of PSII RC donor side. Obtained results indicate that P. zeylanica plants acclimate to lead toxicity by Pb accumulation in roots and, depending on Pb concentration, by adjusting their photosynthetic apparatus via the activation of alternative (cyclic and pseudocyclic) electron transport pathways.

Comparative analysis of epigenetic inhibitors reveals different degrees of interference with transcriptional gene silencing and induction of DNA damage.

Repetitive DNA sequences and some genes are epigenetically repressed by transcriptional gene silencing (TGS). When genetic mutants are not available or problematic to use, TGS can be suppressed by chemical inhibitors. However, informed use of epigenetic inhibitors is partially hampered by the absence of any systematic comparison. In addition, there is emerging evidence that epigenetic inhibitors cause genomic instability, but the nature of this damage and its repair remain unclear. To bridge these gaps, we compared the effects of 5-azacytidine (AC), 2'-deoxy-5-azacytidine (DAC), zebularine and 3-deazaneplanocin A (DZNep) on TGS and DNA damage repair. The most effective inhibitor of TGS was DAC, followed by DZNep, zebularine and AC. We confirmed that all inhibitors induce DNA damage and suggest that this damage is repaired by multiple pathways with a critical role of homologous recombination and of the SMC5/6 complex. A strong positive link between the degree of cytidine analog-induced DNA demethylation and the amount of DNA damage suggests that DNA damage is an integral part of cytidine analog-induced DNA demethylation. This helps us to understand the function of DNA methylation in plants and opens the possibility of using epigenetic inhibitors in biotechnology.

Is a blue-red light a good elicitor of phenolic compounds in the family Droseraceae? A comparative study.

Plants from the family Droseraceae, especially Drosera sp. and Dionaea sp., are naturally rich in phenolic derivatives such as plumbagin, among others. Plumbagin is known both for its pharmacological significance and its protective properties against light stress. Light stress - high light intensity or/and light spectral composition - activates plants' response mechanisms including, among others, hormonal (salicylic acid, jasmonic acid) pathways and secondary metabolite (phenolic compounds, proline) pathways. Short-wavelength radiation, due to its high energy, will induce the synthesis of protective secondary metabolites, including those with pharmaceutical properties. The aim of the study was to describe and compare acclimation strategies of Drosera peltata and Dionaea muscipula to blue-red light in the context of phenolic compound accumulation, and salicylic acid, jasmonic acid and proline synthesis. For the first time, differences in the responses of D. muscipula and D. peltata to blue-red light (in the ratio 6:1) were established. In Dionaea sp., it was associated with the use of redox equivalents (in particular, plastoquinone pool) for the synthesis of primary metabolites used in the process of growth and development. In Drosera sp., a rapid adjustment of redox state led to the synthesis of secondary metabolites, constituting a reservoir of carbon skeletons and allowing for a quick defence response to stress factors. In both species, blue-red light did not induce the jasmonic acid pathway. However, the salicylic acid pathway was induced as an alternative to the phenolic compound synthesis pathway. Nevertheless, the applied blue-red light was not an effective elicitor of phenolic compounds in the plants examined.