Copper-induced oxidative stress and antioxidant defence in Arabidopsis thaliana

Content of reactive oxygen species (ROS): O2*-, H2O2 and OH* as well as activities of antioxidant enzymes: superoxide dismutase (SOD), guaiacol peroxidase (POX) and catalase (CAT) were studied in leaves of Arabidopsis thaliana ecotype Columbia, treated with Cu excess (0, 5, 25, 30, 50, 75, 100, 150 and 300 microM). After 7 days of Cu action ROS content and the activity of SOD and POX increased, while CAT activity decreased in comparison with control. Activities of SOD, POX and CAT were correlated both with Cu concentration (0-75 microM) in the growth medium and with OH* content in leaves. Close correlation was also found between OH* content and Cu concentration. Oxidative stress in A. thaliana under Cu treatment expressed in elevated content of O2*-, H2O2 and OH* in leaves. To overcome it very active the dismutase- and peroxidase-related (and not catalase-related, as in other plants) ROS scavenging system operated in A. thaliana. Visual symptoms of phytotoxicity: chlorosis, necrosis and violet colouring of leaves as well as a reduction of shoot biomass occurred in plants.     

Intraspecific hybrids of Arabidopsis thaliana revealed no gross alterations in endopolyploidy, DNA methylation, histone modifications and transcript levels

Arabidopsis accessions Col-0 and C24 and their reciprocal hybrids were employed as a model system to investigate the potential relationship between changes in DNA methylation, chromatin structure, endopolyploidization and gene expression in heterotic genotypes. Nucleolus size, endopolyploidization level and distribution of DNA and histone H3 methylation at the microscopic level does not differ between parents and their hybrids. Methylation sensitive amplified polymorphism revealed a largely constant pattern of DNA methylation (97% of signals analyzed) after intraspecific crosses. The parental expression profile of selected genes was maintained in hybrid offspring. No correlation was found between expression pattern and DNA methylation levels at restriction sites within 5′ regulatory regions. Thus, the results revealed only minor changes of chromatin properties and other nuclear features in response to intraspecific hybridization in Arabidopsis thaliana.     

Overproduction of Arabidopsis thaliana FeSOD confers oxidative stress tolerance to transgenic maize.

Transgenic maize (Zea mays L.) plants have been generated by particle gun bombardment that overproduce an Arabidopsis thaliana iron superoxide dismutase (FeSOD). To target this enzyme into chloroplasts, the mature Fesod coding sequence was fused to a chloroplast transit peptide from a pea ribulose-1,5-bisphosphate carboxylase gene. Expression of the chimeric gene was driven by the CaMV 35S promoter. Growth characteristics and in vitro oxidative stress tolerance of transgenic lines grown in control and chilling temperatures were evaluated. The transgenic line with the highest transgenic FeSOD activities had enhanced tolerance toward methyl viologen and had increased growth rates.     

Transgenerational response to stress in Arabidopsis thaliana

Plants exposed to stress pass the memory of exposure to stress to the progeny. Previously, we showed that the phenomenon of transgenerational memory of stress is of epigenetic nature and depends on the function of Dicer-like (DCL) 2 and DCL3 proteins. Here, we discuss a possible role of DNA methylation and function of small RNAs in establishing and maintaining transgenerational responses to stress. Our new data report that memory of stress is passed to the progeny predominantly through the female rather than male gamete. Possible evolutionary advantages of this mechanism are also discussed.Key words: transgenerational response to stress, Arabidopsis thaliana, maternal inheritance, methylation changes, homologous recombination frequency, genome instability, adaptive response, dcl2, dcl3Plants are sedentary organisms and thus can not respond to rapidly changing growth conditions by escaping to new environments as animals usually do. Moreover, since seed dispersal is rather limited in the vast majority of plants, the progeny is very likely to grow under the same environmental growth conditions as its parents did. The memory of pre-existing growth conditions can be advantageous for plant survival. The environmental experience of parents can be recorded in the form of induced epigenetic modifications that occur in somatic cell lineages. The very late, almost at the end of plant development, separation of germline cells from somatic tissues enables incorporation of acquired epigenetic changes in the gametes. Indeed, previous reports suggested that the progeny of exposed plants might have an advantage while growing in the same environment as its parents.^1–3 Despite a growing number of experimental evidences that support the existence of the phenomenon of memory of stress, the data on adaptive changes in the progeny of stressed plants are scarce.Parental exposure to stress may not only lead to adaptive effects in progeny but also introduce a certain degree of changes in genome stability.^4–9 Our early report showed that the progeny of tobacco plants infected with tobacco mosaic virus had an increased meiotic recombination frequency.⁸ A more recent report demonstrated that these progeny plants had a higher frequency of rearrangements at the loci carrying the homology to N-gene-like R-gene loci, allowing speculations about a possible role of these rearrangements in pathogen resistance evolution.⁹ Similarly, a study of Molinier et al. (2006) showed that the progeny of plants exposed to UVC or flagellin had an increased frequency of somatic homologous recombination events (HRF).⁴ The authors demonstrated that an increase in HRF triggered by a single exposure to UVC was maintained for five consecutive generations in the absence of stress. In contrast, our most recent reports demonstrated that maintaining an increase in HRF caused by ancestral exposure to heat, cold, flood, UVC or salt required exposure to stress in subsequent generations: if F1 plants were propagated for one more generation without stress, the effect diminished and HRF returned back to the level observed in the progeny of untreated plants.^6,7 This scenario seems to be more probable from an evolutionary point of view. Within a given environmental niche, plants establish certain genetic and epigenetic traits needed to cope with the expected growth conditions. Drastic environmental changes or new unusual stresses may trigger a cascade of gene expression changes in attempt to survive and adapt to new conditions. Some of these potentially advantageous changes are most probably recorded in the form of DNA methylation and chromatin modifications and are passed to progeny as memory of stress exposure.It can be further hypothesized that if these new environmental conditions are no longer present during the lifespan of future generations, the newly established methylation patterns and chromatin organization will return to the original epigenetic landscape that was the most adequate fit for this environmental niche. If the same new stresses occur in consecutive generations, the newly established epigenetic changes will be maintained and possibly stabilized after many generations of exposure.     

Metabolic recovery of Arabidopsis thaliana roots following cessation of oxidative stress

To cope with the various environmental stresses resulting in reactive oxygen species (ROS) production plant metabolism is known to be altered specifically under different stresses. After overcoming the stress the metabolism should be reconfigured to recover basal operation however knowledge concerning how this is achieved is cursory. To investigate the metabolic recovery of roots following oxidative stress, changes in metabolite abundance and carbon flow were analysed. Arabidopsis roots were treated by menadione to elicit oxidative stress. Roots were fed with ¹³C labelled glucose and the redistribution of isotope was determined in order to study carbon flow. The label redistribution through many pathways such as glycolysis, the tricarboxylic acid (TCA) cycle and amino acid metabolism were reduced under oxidative stress. After menadione removal many of the stress-related changes reverted back to basal levels. Decreases in amounts of hexose phosphates, malate, 2-oxoglutarate, glutamate and aspartate were fully recovered or even increased to above the control level. However, some metabolites such as pentose phosphates and citrate did not recover but maintained their levels or even increased further. The alteration in label redistribution largely correlated with that in metabolite abundance. Glycolytic carbon flow reverted to the control level only 18 h after menadione removal although the TCA cycle and some amino acids such as aspartate and glutamate took longer to recover. Taken together, plant root metabolism was demonstrated to be able to overcome menadione-induced oxidative stress with the differential time period required by independent pathways suggestive of the involvement of pathway specific regulatory processes.     

Response to biotic and oxidative stress in Arabidopsis thaliana: analysis of variably phosphorylated proteins

Protein phosphorylation plays a pivotal role in the regulation of many cellular events; increasing evidences indicate that this post-translational modification is involved in plant response to various abiotic and biotic stresses. Since phosphorylated proteins may be present at low abundance, enrichment methods are generally required for their analysis. We here describe the quantitative changes of phosphoproteins present in Arabidopsis thaliana leaves after challenging with elicitors or treatments mimicking biotic stresses, which stimulate basal resistance responses, or oxidative stress. Phosphoproteins from elicited and control plants were enriched by means of metal oxide affinity chromatography and resolved by 2D electrophoresis. A comparison of the resulting proteomic maps highlighted phosphoproteins showing quantitative variations induced by elicitor treatment; these components were identified by MALDI-TOF peptide mass fingerprinting and/or nanoLC-ESI-LIT-MS/MS experiments. In total, 97 differential spots, representing 75 unique candidate phosphoproteins, were characterized. They are representative of different protein functional groups, such as energy and carbon metabolism, response to oxidative and abiotic stresses, defense, protein synthesis, RNA processing and cell signaling. Ascertained protein phosphorylation found a positive confirmation in available Arabidopsis phosphoproteome database. The role of each identified phosphoprotein is here discussed in relation to plant defense mechanisms. Our results suggest a partial overlapping of the responses to different treatments, as well as a communication with key cellular functions by imposed stresses.     

多环芳烃荧蒽诱导拟南芥氧化胁迫

选用模式植物拟南芥为材料,以四环的多环芳烃（PAHs）荧蒽为研究对象,从植物对非生物胁迫响应紧密相关的抗氧化酶及膜保护系统的变化入手,研究了植物对多环芳烃胁迫的生理响应.结果表明：荧蒽胁迫下拟南芥经历了氧化胁迫和膜脂过氧化过程.0.75 mmol·L^-1的荧蒽使拟南芥光合作用过程受到抑制;1.00 mmol·L^-1的荧蒽使拟南芥丙二醛(MDA)含量极显著增加, 抗坏血酸过氧化物酶(APX)活性极显著下降, 膜脂过氧化作用加剧,1.25 mmol·L^-1的荧蒽使拟南芥过氧化物酶(POD)活性极显著下降,H₂O₂在细胞内累积,拟南芥明显受害.     

Gravitropism in the starch excess mutant of Arabidopsis thaliana

Amyloplasts are hypothesized to play a key role in the cellular mechanisms of gravity perception in plants. While previous studies have examined the effects of starch deficiency on gravitropic sensitivity, in this paper, we report on gravitropism in plants with a greater amount of starch relative to the normal wild type. Thus, we have studied the sex1 (starch excess) mutant of Arabidopsis thaliana, which accumulates extra starch because it is defective in a protein involved in the regulation of starch mobilization. Compared to the wild type (WT), sex1 seedlings contained excess starch in cotyledons, hypocotyls, the root-hypocotyl transition zone, the body of the root, root hairs, and in peripheral rootcap cells. Sedimented amyloplasts were found in both the WT and in sex1 in the rootcap columella and in the endodermis of stems, hypocotyls, and petioles. In roots, the starch content and amyloplast sedimentation in central columella cells and the gravitropic sensitivity were comparable in sex1 and the WT. However, in hypocotyls, the sex1 mutant was much more sensitive to gravity during light-grown conditions compared to the WT. This difference was correlated to a major difference in size of plastids in gravity-perceiving endodermal cells between the two genotypes (i.e., sex1 amyloplasts were twice as big). These results are consistent with the hypothesis that only very large changes in starch content relative to the WT affect gravitropic sensitivity, thus indicating that wild-type sensing is not saturated.     

Ultraviolet-B exposure leads to up-regulation of senescence-associated genes in Arabidopsis thaliana.

Exposure to UV-B radiation resulted in a loss of chlorophyll and an increase in lipid damage in a similar manner to that induced during natural senescence. In addition, exposure to UV-B led to the induction of a number of genes associated with senescence (SAG12, 13, 14, and 17). These results show, for the first time, that exposure to UV-B can lead to cellular decline through active and regulated processes involving many genes also associated with natural senescence.     

Salt and osmotic stress cause rapid increases in Arabidopsis thaliana cGMP levels

A guanylyl cyclase has been recently identified in Arabidopsis but, despite the use of pharmacological inhibitors to infer roles of the second messenger 3',5'-cyclic guanosine monophosphate (cGMP), very few measurements of actual cGMP levels in plants are available. Here, we demonstrate that cGMP levels in Arabidopsis seedlings increase rapidly (< or =5 s) and to different degrees after salt and osmotic stress, and that the increases are prevented by treatment with LY, an inhibitor of soluble guanylyl cyclases. In addition, we provide evidence to suggest that salt stress activates two cGMP signalling pathways - an osmotic, calcium-independent pathway and an ionic, calcium-dependent pathway.     

Study of oxidative stress related responses induced in Arabidopsis thaliana following mixed exposure to uranium and cadmium

In this study, toxicity effects in plants of uranium in a binary pollution condition were investigated by studying biological responses and unraveling oxidative stress related mechanisms in Arabidopsis thaliana seedlings, grown on hydroponics and exposed for 3 days to 10 μM uranium in combination with 5 μM cadmium. While uranium mostly accumulated in the roots with very low root-to-shoot transport, cadmium was taken up less by the roots but showed higher translocation to the shoots. Under mixed exposure, cadmium influenced uranium uptake highly but not the other way round resulting in a doubled uranium concentration in the roots. Under our mixed exposure conditions, it is clear that micronutrient concentrations in the roots are strongly influenced by addition of cadmium as a second stressor, while leaf macronutrient concentrations are mostly influenced by uranium. Oxidative stress related responses are highly affected by cadmium while uranium influence is more limited. Hereby, an important role was attributed to the ascorbate redox balance together with glutathione as both metabolites, but more explicitly for ascorbate, increased their reduced form, indicating an important defense and regulatory function. While for roots, based on an increase in FSD1 gene expression, oxidative stress was suggested to be superoxide induced, in leaves on the other hand, hydrogen peroxide related genes were mostly altered.     

Enhanced oxidative stress in the ethylene-insensitive (ein3-1) mutant of Arabidopsis thaliana exposed to salt stress

To better understand the role of ethylene signaling in plant stress tolerance, salt-induced changes in gene expression levels of ethylene biosynthesis, perception and signaling genes were measured in Arabidopsis thaliana plants exposed to 15 days of salinity. Among the genes analyzed, EIN3 showed the highest expression level increase under salt stress, suggesting a key role for this ethylene-signaling component in response to salt stress. Therefore, we analyzed the salt stress response over 15 days (by adding 100 mM NaCl to the nutrient solution) in the ein3-1 mutant compared to the wild-type (Col-0) in terms of growth, oxidative stress markers (lipid peroxidation, foliar pigments and low-molecular-weight antioxidants) and levels of growth- and stress-related phytohormones (including cytokinins, auxins, gibberellins, abscisic acid, jasmonic acid and salicylic acid). The ein3-1 mutant grew similarly to wild-type plants both under control and salt stress conditions, which was associated with a differential time course evolution in the levels of the cytokinins zeatin and zeatin riboside, and the auxin indole-3-acetic acid between the ein3-1 mutant and the wild-type. Despite showing no signs of physiological deterioration under salt stress (in terms of rosette biomass, leaf water and pigment contents, and PSII efficiency) the ein3-1 mutant showed enhanced lipid peroxidation under salt stress, as indicated by 2.4-fold increase in both malondialdehyde and jasmonic acid contents compared to the wild-type. We conclude that, at moderate doses of salinity, partial insensitivity to ethylene might be compensated by changes in endogenous levels of other phytohormones and lipid peroxidation-derived signals in the ein3-1 mutant exposed to salt stress, but at the same time, this mutant shows higher oxidative stress under salinity than the wild-type.