Vitamin E protects against photoinhibition and photooxidative stress in Arabidopsis thaliana

Vitamin E is considered a major antioxidant in biomembranes, but little evidence exists for this function in plants under photooxidative stress. Leaf discs of two vitamin E mutants, a tocopherol cyclase mutant (vte1) and a homogentisate phytyl transferase mutant (vte2), were exposed to high light stress at low temperature, which resulted in bleaching and lipid photodestruction. However, this was not observed in whole plants exposed to long-term high light stress, unless the stress conditions were extreme (very low temperature and very high light), suggesting compensatory mechanisms for vitamin E deficiency under physiological conditions. We identified two such mechanisms: nonphotochemical energy dissipation (NPQ) in photosystem II (PSII) and synthesis of zeaxanthin. Inhibition of NPQ in the double mutant vte1 npq4 led to a marked photoinhibition of PSII, suggesting protection of PSII by tocopherols. vte1 plants accumulated more zeaxanthin in high light than the wild type, and inhibiting zeaxanthin synthesis in the vte1 npq1 double mutant resulted in PSII photoinhibition accompanied by extensive oxidation of lipids and pigments. The single mutants npq1, npq4, vte2, and vte1 showed little sensitivity to the stress treatments. We conclude that, in cooperation with the xanthophyll cycle, vitamin E fulfills at least two different functions in chloroplasts at the two major sites of singlet oxygen production: preserving PSII from photoinactivation and protecting membrane lipids from photooxidation.     

Early photosynthetic response of Arabidopsis thaliana to temperature and salt stress conditions

Temperature changes and salt accumulation are among the most common abiotic factors affecting plants in agricultural and natural ecosystems. The different responses of plants to these factors have been widely investigated in previous works. However, detailed mechanism of the early photosynthetic response (first 24 h) has been poorly studied. The aim of the work was to monitor the early response of adult Arabidopsis thaliana plants exposed to different thermal (cold and heat) and salt conditions. Detailed evaluation of the efficiency of photosystem II was done, and the various routes of energy output as well as measurements of the contents of H₂O₂, proline, and photosynthetic pigments at different times during the first 24 h of treatment were examined. The conditions used in the study were those that caused a weak stress with time of exposure. Cold-treated plants showed the most continuous inhibitory effect on photosynthetic activity, with a fast metabolic slowdown (reduced PSII efficiency and decreased pigment contents), although they also demonstrated clear acclimation responses (increased heat dissipation and protein content). Heat-treated plants showed a late but stronger effect on photosynthesis with significantly increased quantum yield of nonregulated energy dissipation (??_NO) and H₂O₂ content at the last measurements. Finally, salt-induced oxidative stress (increased H₂O₂ content), decreased PSII efficiency and pigment content.     

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.     

The Arabidopsis thaliana sulfiredoxin is a plastidic cysteine-sulfinic acid reductase involved in the photooxidative stress response

The 2-cysteine peroxiredoxins (2-Cys-Prxs) are antioxidants that reduce peroxides through a thiol-based mechanism. During catalysis, these ubiquitous enzymes are occasionally inactivated by the substrate-dependent oxidation of the catalytic cysteine to the sulfinic acid (-SO2H) form, and are reactivated by reduction by sulfiredoxin (Srx), an enzyme recently identified in yeast and in mammal cells. In plants, 2-Cys-Prxs constitute the most abundant Prxs and are located in chloroplasts. Here we have characterized the unique Srx gene in Arabidopsis thaliana (AtSrx) from a functional point of view, and analyzed the phenotype of two AtSrx knockout (AtSrx-) mutant lines. AtSrx is a chloroplastic enzyme displaying sulfinic acid reductase activity, as shown by the ability of the recombinant AtSrx to reduce the overoxidized 2-Cys-Prx form in vitro, and by the accumulation of the overoxidized Prx in mutant lines lacking Srx in vivo. Furthermore, AtSrx mutants exhibit an increased tolerance to photooxidative stress generated by high light combined with low temperature. These data establish that, as in yeast and in mammals, plant 2-Cys-Prxs are subject to substrate-mediated inactivation reversed by Srx, and suggest that the 2-Cys-Prx redox status and sulfiredoxin are parts of a signaling mechanism participating in plant responses to oxidative stress.     

Short-term down-regulation of zeaxanthin epoxidation in Arabidopsis thaliana in response to photo-oxidative stress conditions

The epoxidation of zeaxanthin (Zx) to violaxanthin after exposure to different light stress conditions has been studied in Arabidopsis (Arabidopsis thaliana). Formation of Zx was induced by illumination of intact leaves for up to 8 h at different light intensities and temperatures. The kinetics of epoxidation was found to be gradually retarded with increasing light stress during pre-illumination, indicating a gradual down-regulation of the Zx epoxidase activity. Retardation of the epoxidation rates by a factor of up to 10 was inducible either by increasing the light intensity or by extending the illumination time or by decreasing the temperature during pre-illumination. The retardation of the epoxidation kinetics was correlated with a decrease of the PSII quantum efficiency after the pre-illumination treatment. Experiments with the stn7/stn8 mutant of Arabidopsis indicated that the thylakoid protein kinases STN7 and STN8, which are required for the phosphorylation of PSII proteins, are not involved in the short-term down-regulation of Zx epoxidation. However, the retardation of Zx epoxidation was maintained in thylakoids isolated from pre-illuminated leaves, indicating that a direct modification of the Zx epoxidase is most likely involved in the light-induced down-regulation.     

Singlet oxygen scavenging activity of tocopherol and plastochromanol in Arabidopsis thaliana: relevance to photooxidative stress

In the present study, singlet oxygen (¹O₂) scavenging activity of tocopherol and plastochromanol was examined in tocopherol cyclase‐deficient mutant (vte1) of Arabidopsis thaliana lacking both tocopherol and plastochromanol. It is demonstrated here that suppression of tocopherol and plastochromanol synthesis in chloroplasts isolated from vte1 Arabidopsis plants enhanced ¹O₂ formation under high light illumination as monitored by electron paramagnetic resonance spin‐trapping spectroscopy. The exposure of vte1 Arabidopsis plants to high light resulted in the formation of secondary lipid peroxidation product malondialdehyde as determined by high‐pressure liquid chromatography. Furthermore, it is shown here that the imaging of ultra‐weak photon emission known to reflect oxidation of lipids was unambiguously higher in vte1 Arabidopsis plants. Our results indicate that tocopherol and plastochromanol act as efficient ¹O₂ scavengers and protect effectively lipids against photooxidative damage in Arabidopsis plants.     

N-glucosyltransferase UGT76C2 is involved in cytokinin homeostasis and cytokinin response in Arabidopsis thaliana

Cytokinins are a class of phytohormones that play a crucial role in plant growth and development. The gene UGT76C2 encoding cytokinin N-glucosyltransferase of Arabidopsis thaliana has been previously identified. To determine the in planta role of UGT76C2 in cytokinin metabolism and response, we analyzed the phenotypes of its loss-of-function mutant (ugt76c2) and its overexpressors. The accumulation level of the cytokinin N-glucosides was significantly decreased in ugt76c2, but substantially increased in UGT76C2 overexpressors compared with the wild type. When treated with exogenously applied cytokinin, ugt76c2 showed more sensitivity and UGT76C2 overexpressors showed less sensitivity to cytokinin in primary root elongation, lateral root formation, Chl retention and anthocyanin accumulation. Under normal growth conditions ugt76c2 had smaller seeds than the wild type, with accompanying lowered levels of active and N-glucosylated cytokinin forms. The expression levels of cytokinin-related genes such as AHK2, AHK3, ARR1, IPT5 and CKX3 were changed in ugt76c2, suggesting homeostatic control of cytokinin activity. Studies of spatiotemporal expression patterns showed that UGT76C2 was expressed at a relatively higher level in the seedling and developing seed. In their entirety, our data, based mainly on this comparison and opposite phenotypes of knockout and overexpressors, strongly suggest that UGT76C2 is involved in cytokinin homeostasis and cytokinin response in planta through cytokinin N-glucosylation.     

Stomatal response to vapour pressure deficit and the effect of plant water stress

Abstract The dynamic response of stomata to changes in atmospheric humidity was investigated in Fragaria × ananassa Duch., Picea engelmannii Parry, and Pseudotsuga menziesii (Mirb.) Franco; and the effect of water stress on this response was determined in Pseudotsuga menziesii. The plants were rotated through three regimes of ambient temperature and vapour pressure deficit: 35°C–3. 5kPa, 35°C–0. 5 kPa, and 20°C–1. 5kPa. Branch and leaflet conductance were measured with a steady-state porometer, first at ambient vapour pressure deficit and then at one of four treatment conditions achieved by increasing or decreasing vapour pressure within the porometer cuvette. All three species showed similar stomatal response: enhanced conductance at low vapour pressure deficit and depressed conductance at high vapour pressure deficit. Engelmann spruce was more sensitive than Douglas fir and strawberry. Plant water status significantly altered stomatal response to vapour pressure deficit. The relationship of conductance of xylem water potential was linear under ambient conditions but became curvilinear when conductance was measured above and below ambient vapour pressure deficit. Between ?0. 5 MPa and ?2. 0 MPa xylem water potential, the stomata were sensitive to vapour pressure deficit, but below ? 2. 0 MPa, the sensitivity decreased.     

Growth of Arabidopsis thaliana seedlings under water deficit studied by control of water potential in nutrient-agar media

We have characterized the growth responses of Arabidopsis thaliana seedlings to water deficit. To manipulate the water potential, we developed a method whereby the nutrient-agar medium could be supplemented with polyethylene glycol (PEG 8000); PEG was introduced into gelled media by diffusion, which produced media with water potential as low as -1.6 MPa. For dark-grown plants, hypocotyl growth had a hyperbolic dependence on water potential, and was virtually stopped by -1 MPa. In contrast, primary root elongation was stimulated by moderate deficit and even at -1.6 MPa was not significantly less than the control. That these results did not depend on a direct effect of PEG was attested by obtaining indistinguishable results when a dialysis membrane impermeable to PEG was placed between the medium and the seedlings. For light-grown seedlings, moderate deficit also stimulated primary root elongation and severe deficit reduced elongation only partially. These changes in elongation were paralleled by changes in root system dry weight. At moderate deficit, lateral root elongation and initiation were unaffected and at higher stress levels both were inhibited. Primary root diameter increased steadily with time in well-watered controls and under water deficit increased transiently before stabilizing at a diameter that was inversely proportional to the deficit. Along with stimulated primary root elongation, moderate water deficit also stimulated the rate of cell production. Thus, A. thaliana responds to water deficit vigorously, which enhances its use as a model to uncover mechanisms underlying plant responses to water deficit.     

Monitoring genome-wide changes in gene expression in response to endogenous cytokinin reveals targets in Arabidopsis thaliana

Cytokinins have been implicated in developmental and growth processes in plants including cell division, chloroplast biogenesis, shoot meristem initiation and senescence. The regulation of these processes requires changes in cytokinin-responsive gene expression. Here, we induced the expression of a bacterial isopentenyl transferase gene, IPT, in transgenic Arabidopsis thaliana seedlings to study the regulation of genome-wide gene expression in response to endogenous cytokinin. Using MPSS (massively parallel signature sequencing) we identified 823 and 917 genes that were up- and downregulated, respectively, following 24 h of IPT induction. When comparing the response to cytokinin after 6 and 24 h, we identified different clusters of genes showing a similar course of regulation. Our study provides researchers with the opportunity to rapidly assess whether genes of interest are regulated by cytokinins.     

Ascorbate peroxidase 1 plays a key role in the response of Arabidopsis thaliana to stress combination

Within their natural habitat plants are subjected to a combination of different abiotic stresses, each with the potential to exacerbate the damage caused by the others. One of the most devastating stress combinations for crop productivity, which frequently occurs in the field, is drought and heat stress. In this study we conducted proteomic and metabolic analysis of Arabidopsis thaliana plants subjected to a combination of drought and heat stress. We identified 45 different proteins that specifically accumulated in Arabidopsis in response to the stress combination. These included enzymes involved in reactive oxygen detoxification, malate metabolism, and the Calvin cycle. The accumulation of malic enzyme during the combined stress corresponded with enhanced malic enzyme activity, a decrease in malic acid, and lower amounts of oxaloacetate, suggesting that malate metabolism plays an important role in the response of Arabidopsis to the stress combination. Cytosolic ascorbate peroxidase 1 (APX1) protein and mRNA accumulated during the stress combination. When exposed to heat stress combined with drought, an APX1-deficient mutant (apx1) accumulated more hydrogen peroxide and was significantly more sensitive to the stress combination than wild type. In contrast, mutants deficient in thylakoid or stromal/mitochondrial APXs were not more sensitive to the stress combination than apx1 or wild type. Our findings suggest that cytosolic APX1 plays a key role in the acclimation of plants to a combination of drought and heat stress.