Transgenic tobacco plants expressing antisense ferredoxin-NADP(H) reductase transcripts display increased susceptibility to photo-oxidative damage

Ferredoxin-NADP(H) reductase (FNR) catalyses the final step of the photosynthetic electron transport in chloroplasts. Using an antisense RNA strategy to reduce expression of this flavoenzyme in transgenic tobacco plants, it has been demonstrated that FNR mediates a rate-limiting step of photosynthesis under both limiting and saturating light conditions. Here, we show that these FNR-deficient plants are abnormally prone to photo-oxidative injury. When grown under autotrophic conditions for 3 weeks, specimens with 20-40% extant reductase undergo leaf bleaching, lipid peroxidation and membrane damage. The magnitude of the effect was proportional to the light intensity and to the extent of FNR depletion, and was accompanied by morphological changes involving accumulation of aberrant plastids with defective thylakoid stacking. Damage was initially confined to chloroplast membranes, whereas Rubisco and other stromal proteins began to decline only after several weeks of autotrophic growth, paralleled by partial recovery of NADPH levels. Exposure of the transgenic plants to moderately high irradiation resulted in rapid loss of photosynthetic capacity and accumulation of singlet oxygen in leaves. The collected results suggest that the extensive photo-oxidative damage sustained by plants impaired in FNR expression was caused by singlet oxygen building up to toxic levels in these tissues, as a direct consequence of the over-reduction of the electron transport chain in FNR-deficient chloroplasts.     

Arabidopsis mutants deficient in diacylglycerol acyltransferase display increased sensitivity to abscisic acid,sugars, and osmotic stress during germination and seedling development

Arabidopsis seeds store triacylglycerol (TAG) as the major carbon reserve, which is used to support postgerminative seedling growth. Diacylglycerol acyltransferase (DGAT) catalyzes the final step in TAG synthesis, and two isoforms of DGAT have previously been identified in Arabidopsis. It has been shown that DGAT1 plays an important role in seed development because Arabidopsis with mutations at the TAG1 locus accumulate less seed oil. There is also evidence showing that DGAT1 is active after seed germination. The aim of this study is to investigate the effect of mutations of DGAT1 on postembryonic development in Arabidopsis. We carried out detailed analyses of two tag1 mutants in different ecotypic backgrounds of Arabidopsis. Results show that during germination and seedling growth, seed storage TAG degradation was not affected in the tag1 mutants. However, sugar content of the mutant seedlings is altered, and activities of the hexokinases are significantly increased in the tag1 mutant seedlings. The tag1 mutants are also more sensitive to abscisic acid, glucose, and osmotic strength of the medium in germination and seedling growth.     

Down syndrome fibroblasts and mouse Prep1-overexpressing cells display increased sensitivity to genotoxic stress

PREP1 (PKNOX1) maps in the Down syndrome (DS) critical region of chromosome 21, is overexpressed in some DS tissues and might be involved in the DS phenotype. By using fibroblasts from DS patients and by overexpressing Prep1 in F9 teratocarcinoma and Prep1^i/i MEF to single out the role of the protein, we report that excess Prep1 increases the sensitivity of cells to genotoxic stress and the extent of the apoptosis directly correlates with the level of Prep1. The apoptotic response of Prep1-overexpressing cells is mediated by the pro-apoptotic p53 protein that we show is a direct target of Prep1, as its depletion reverts the apoptotic phenotype. The induction of p53 overcomes the anti-apoptotic role of Bcl-X_L, previously shown to be also a Prep1 target, the levels of which are increased in Prep1-overexpressing cells as well. Our results provide a rationale for the involvement of PREP1 in the apoptotic phenotype of DS tissues and indicate that differences in Prep1 level can have drastic effects.     

Singlet oxygen and photo-oxidative stress management in plants and algae

Photosynthetic organisms constantly face the threat of photo-oxidative stress from fluctuating light conditions and environmental stress. Plants and algae have developed an array of defences to protect the chloroplast from reactive oxygen species. Genetic and physiological studies have shown that antioxidant responses are important to high-light acclimation, both by directly scavenging or quenching reactive oxygen intermediates and by contributing reducing power for alternative electron transport pathways and excess energy dissipation. At present, the signalling events leading to up-regulation of antioxidant defences in high light remain a mystery. Recent advances toward understanding acclimation to oxidative stress in both photosynthetic and non-photosynthetic model organisms may illuminate how plants and algae respond to high-light stress. Although the role of hydrogen peroxide in high-light acclimation has been investigated, less is known about responses to singlet oxygen, a form of reactive oxygen that poses a significant threat specifically to photosynthetic organisms. This review will discuss some intriguing new findings in that area, focusing on recent findings regarding the nature of singlet-oxygen responses in the chloroplast.     

Genetic analysis of tolerance to photo-oxidative stress induced by high light in winter wheat （Triticum aestivum L.）

High light induced photooxidation (HLIP) usually leads to leaf premature senescence and causes great yield loss in winter wheat. In order to explore the genetic control of wheat tolerance to HLIP stress, a quantitative trait loci (QTL) analysis was conducted on a set of doubled haploid population, derived from two winter wheat cultivars. Actual values of chlorophyll content (Chl), minimum fluorescence level (Fo), maximum fluorescence level (Fm), and the maximum quantum efficiency of photosystem Ⅱ (Fv/Fm) under both HLIP and non-stress conditions as well as the ratios of HLIP to non-stress were evaluated. HLIP considerably reduced Chl, Fm, and Fv/Fm, but increased Fo, compared with that under non-stress condition. A total of 27, 16, and 28 QTLs were associated with the investigated traits under HLIP and non-stress and the ratios of HLIP to non-stress, respectively. Most of the QTLs for the ratios of HLIP to non-stress collocated or nearly linked with those detected under HLIP condition. HLIP-induced QTLs were mapped on 15 chromosomes, involving in 1A, 1B, 1D,2A, 2B, 2D, 3A, 3B, 4A, 4D, 5B, 6A, 6B, 7A, and 7D while those expressed under non-stress condition involved in nine chromosomes, including 1B, 1D, 2A, 2B, 3B, 4A, 5A, 5B, and 7A. The expression patterns of QTLs under HLIP condition were different from that under non-stress condition except for six loci on five chromosomes. The phenotypic variance explained by individual QTL ranged from 5.0% to 19.7% under HLIP, 8.3% to 20.8% under non-stress, and 4.9% to 20.2% for the ratios of HLIP to non-stress, respectively. Some markers, for example,Xgwm192 and WMC331 on 4D regulating Chl, Fo, Fm, and Fv/Fm under HLIP condition, might be used in marker assistant selection.     

COE 1 and GUN1 regulate the adaptation of plants to high light stress

In order to withstand high light (HL) stress, plants have evolved both short-term defense and repair mechanisms and long-term acclimation responses. At present, however, the underlying signaling events and molecular mechanisms are still poorly understood. Analysis of the mutants coe1, coe1 gun1 double mutant and oeGUN1coe1 revealed increased sensitivity to HL stress as compared to wild type (WT), with oeGUN1 coe1 plants displaying the highest sensitivity. Accumulation of FTSH2 protein and degradation of D1 protein during the HL stress were shown to depend on both COE1 and GUN1. Overexpression of COE1 enhanced the induction of FTSH2 and the tolerance to HL stress. These results indicate that the COE1-GUN1 signaling pathway plays an important role in regulating the adaptation of plants to HL.     

Characterization of a rice mutant having an increased susceptibility to light stress at high temperature

The effect of light irradiation at high temperature was investigated in a rice mutant, spl-2 , which is sensitive to solar radiation. Dead spots appeared on the mutant leaves when cultured at a high temperature (40°C) under strong white-light illumination (15 W m⁻²). A similar damage was also observed in the wild-type leaves under the same conditions when the plants were preincubated in the dark for one day. Preillumination with weak light (6 W m⁻²) lessened the irradiation-induced damage in the wild-type. These observations suggest that in rice plants the acclimatization to weak light has a protective effect against strong irradiation at high temperature, and the spl-2 locus participates in the mechanisms of the acclimatization. The action spectrum for the irradiation-induced damage at the high temperature (40°C) in the spl-2 leaves indicated that the maximum damage occurred at around 480 nm and, in a lesser extent, at around 680 nm. Activity of the O⁻₂₋ and H₂O₂₋scavenging enzymes in the spl−2 leaves were almost the same or somewhat higher than those in the wild-type after irradiation with strong white light (15 W m⁻²) at 40°C, whereas the content of ascorbic acid in the spl−2 decreased significantly compared with the wild type.     

Mitochondrial dysfunction and increased sensitivity to excitotoxicity in mice deficient in DNA mismatch repair

The expression profile in the hippocampus of mice lacking one allele of the MutS homologue (Msh2), gene, which is one of the most representative components of the DNA mismatch repair system, was analysed to understand whether defects in the repair or in response to DNA damage could impact significantly on brain function. The overall results suggested a reduction in mitochondrial function as indicated by gene expression analysis, biochemical and behavioural studies. In the hippocampus of Msh2+/- mice, array data, validated by RT-PCR and western blot analysis, showed reduced expression levels of genes for cytochrome c oxidase subunit 2 (CoxII), ATP synthase subunit beta and superoxide dismutase 1. Biochemically, mitochondria from the hippocampus and cortex of these mice show reduced CoxII and increased aconitase activity. Behaviourally, these alterations resulted in mice with increased vulnerability to kainic acid-induced epileptic seizures and hippocampal neuronal loss. These data suggest that lack of an efficient system involved in recognizing and repairing DNA damage may generate a brain mitochondriopathy.     

Growth condition-dependent sensitivity, photodamage and stress response of Chlamydomonas reinhardtii exposed to high light conditions

Different substrate conditions, such as varying CO(2) concentrations or the presence of acetate, strongly influence the efficiency of photosynthesis in Chlamydomonas reinhardtii. Altered photosynthetic efficiencies affect the susceptibility of algae to the deleterious effects of high light stress, such as the production of reactive oxygen species (ROS) and PSII photodamage. In this study, we investigated the effect of high light on C. reinhardtii grown under photomixotrophy, i.e. in the presence of acetate, as well as under photoautotrophic growth conditions with either low or high CO(2) concentrations. Different parameters such as growth rate, chlorophyll bleaching, singlet oxygen generation, PSII photodamage and the total genomic stress response were analyzed. Although showing a similar degree of PSII photodamage, a much stronger singlet oxygen-specific response and a broader general stress response was observed in acetate and high CO(2)-supplemented cells compared with CO(2)-limited cells. These different photooxidative stress responses were correlated with the individual cellular PSII content and probably directly influenced the ROS production during exposure to high light. In addition, growth of high CO(2)-supplemented cells was more susceptible to high light stress compared with cells grown under CO(2) limitation. The growth of acetate-supplemented cultures, on the other hand, was less affected by high light treatment than cultures grown under high CO(2) concentrations, despite the similar cellular stress. This suggests that the production of ATP by mitochondrial acetate respiration protects the cells from the deleterious effects of high light stress, presumably by providing energy for an effective defense.     

Increased target tissue uptake of,and sensitivity to,testosterone in the vitamin B6 deficient rat

Six-week old male rats were maintained for 4 weeks on a vitamin B₆-free diet to cause a moderately severe degree of vitamin b₆ depletion. This led to a significant reduction in the circulating concentration of testosterone in plasma (control = 8.36 ± 1.68, deficient = 2.13 ± 0.54 nmol/1), but had no effect on circulating concentrations of luteinizing hormone, or, in intact males, on the weight of the prostate relative to body weight. In both intact and 24-h castrated animals vitamin b₆ deficiency resulted in a significant increase in the uptake of [³H]testosterone into the prostate, and both increased and prolonged the specific nuclear retention of the steroid, as assessed by the ratio of radioactivity in the nuclear pellet: the high speed supernatant fraction. The results suggest that vitamin b₆ has a function in the action of testosterone (and other steroid hormones), possibly in the recycling of receptors from the nucleus back into the cytosol after initial translocation. Vitamin b₆ deficient animals have either a reduced rate of synthesis of testosterone or an increased rate of metabolic clearance compared with vitamin b₆ supplemented controls, and this appears to be associated with enhanced target organ response to the hormone.     

The protoplasmic viscosity of terrestrial plants and its sensitivity to light

Summary According to earlier investigations, light initiates changes in the displacement of the chloroplasts on centrifugation in bothHelodea andSpirogyra. These changes, which are interpreted as a result of corresponding changes in the viscosity of the protoplasm, are found to occur in terrestrial plants as well.The plants used for the experiments areAchillea millefolium, Brassica napobrassica, Beta vulgaris andRumex acetosella. If the leaves are kept in darkness and saturated moisture for 2–3 days, the protoplasm attains some degree of internal equilibrium, and its fluidity becomes relatively constant. When the leaves are subsequently exposed to light, this equilibrium is disturbed. Even brief illumination (e.g. 100 m.c. for 1 minute) initiates such changes that the displacement of the chloroplasts on centrifugation is alternately increased and decreased. These fluctuations, which seem to be irregular with respect to both duration and amplitude, continue for several hours after cessation of illumination.     

Transgenic tobacco plants overexpressing chloroplastic ferredoxin-NADP(H) reductase display normal rates of photosynthesis and increased tolerance to oxidative stress

Ferredoxin-NADP(H) reductase (FNR) catalyzes the last step of photosynthetic electron transport in chloroplasts, driving electrons from reduced ferredoxin to NADP+. This reaction is rate limiting for photosynthesis under a wide range of illumination conditions, as revealed by analysis of plants transformed with an antisense version of the FNR gene. To investigate whether accumulation of this flavoprotein over wild-type levels could improve photosynthetic efficiency and growth, we generated transgenic tobacco (Nicotiana tabacum) plants expressing a pea (Pisum sativum) FNR targeted to chloroplasts. The alien product distributed between the thylakoid membranes and the chloroplast stroma. Transformants grown at 150 or 700 micromol quanta m(-2) s(-1) displayed wild-type phenotypes regardless of FNR content. Thylakoids isolated from plants with a 5-fold FNR increase over the wild type displayed only moderate stimulation (approximately 20%) in the rates of electron transport from water to NADP+. In contrast, when donors of photosystem I were used to drive NADP+ photoreduction, the activity was 3- to 4-fold higher than the wild-type controls. Plants expressing various levels of FNR (from 1- to 3.6-fold over the wild type) failed to show significant differences in CO2 assimilation rates when assayed over a range of light intensities and CO2 concentrations. Transgenic lines exhibited enhanced tolerance to photooxidative damage and redox-cycling herbicides that propagate reactive oxygen species. The results suggest that photosynthetic electron transport has several rate-limiting steps, with FNR catalyzing just one of them.     

Chilling-induced leaf abscission of Ixora coccinea plants. I. Induction by oxidative stress via increased sensitivity to ethylene

Exposing ixora ( Ixora coccinea ) plants to chilling temperatures (3–9°C for 3 days) resulted in increased leaf abscission, initiated 3 days after transfer to 20°C. Exposure to chilling also induced a 7-fold increase in ethylene production rates of abscission zone (AZ) tissue during the initial 5 h after chilling. The ethylene burst resulted from the high levels of 1-aminocyclopropane-1-carboxylic acid (ACC) accumulated in the AZ during the chilling period. ACC levels following chilling decreased also due to enhanced conjugation to 1-(malonylamino)cyclopropane-1-carboxylic acid (MACC). Treating plants prior to chilling with antioxidants, such as butylated hydroxyanisole (BHA), n -propyl gallate (PG), and vitamin E, significantly reduced chilling-induced leaf abscission. This effect was obtained despite the fact that ethylene production in the treated plants resembled that of chilled plants receiving no BHA. In addition, exposure of plants to ethylene (0.5–10 μl l⁻¹) for 1–3 days significantly enhanced leaf abscission only when they had been pre-chilled. These data imply that chilling-induced leaf abscission was closely correlated with increased sensitivity of the AZ to ethylene rather than with the chilling-induced ethylene burst. Based on the findings that the ethylene action inhibitor, 1-methylcyclopropene (1-MCP), and the antioxidant BHA inhibited both the chilling-induced and the ethylene-enhanced leaf abscission, it is concluded that: (1) although ethylene is essential for chilling-induced abscission, it is not the triggering factor; (2) oxidative processes derived from the chilling stress seem to be the trigger of chilling-induced leaf abscission, operating via increased sensitivity to ethylene.     

In vitro and in vivo activated T cells display increased sensitivity to PGE2.

In this study we report that in vitro activation of T cells increased the cyclic AMP response to subsequent prostaglandin E2 (PGE2) stimulation severalfold per cell. This sensitization of T cells to PGE2-induced cyclic AMP generation was observed when the T cells had been stimulated in vitro for 5 days with either the CD3 monoclonal antibody OKT3, phytohemagglutinin, or the combination phytohemagglutinin plus the phorbol ester PMA. Enhanced cyclic AMP generation following mitogenic activation was seen in response to both PGE2 and forskolin, direct activator of the adenylate cyclase, indicating that the amount of adenylate cyclase had increased during the in vitro activation course. In order to investigate whether various T cell subsets in general and in vivo activated T cells in particular would differ in their susceptibility to PGE2, we isolated CD4+, CD8+, CD4-CD8-, CD4+CD45RO+ ("memory"), and CD4+CD45RA+ ("virgin") T cells and studied PGE2-mediated inhibition of CD3-induced proliferation, as well as cyclic AMP generation in response to PGE2, respectively. We found that CD8+ T cells are more susceptible to PGE2 inhibition and produce more cyclic AMP than CD4+ T cells. Double-negative T cells (enriched for gamma delta T cell receptor positive cells) were found to be sensitive to PGE2 as well. Within the CD4+ T cell population, CD45RO+ ("memory") T cells were significantly more sensitive to PGE2-mediated suppression than CD45RA+ ("virgin") T cells. CD45RO+ cells required a 10-fold lower dose of PGE2 for half-maximum suppression of proliferation. However, no difference in cyclic AMP production could be demonstrated between these two subsets. We propose that substantial heterogeneity exists among peripheral blood T lymphocyte subsets regarding their sensitivity to the immunosuppressive action of PGE2 and that the sensitivity of individual cells changes in the course of an immune response.     

Overexpression of zeaxanthin epoxidase gene enhances the sensitivity of tomato PSII photoinhibition to high light and chilling stress

A tomato ( Lycopersicon esculentum Mill.) zeaxanthin epoxidase gene ( LeZE ) was isolated. The deduced amino acid sequence of LeZE showed high identities with zeaxanthin epoxidase in other plant species. Northern blot analysis showed that the mRNA accumulation of LeZE in the wild-type (WT) was not induced by light and temperature but regulated by the diurnal rhythm. The sense transgenic plants were obtained under the control of the cauliflower mosaic virus 35S promoter (35S-CaMV). Northern and western blot analysis confirmed that sense LeZE was transferred into the tomato genome and overexpressed. The ratio of (A + Z)/(V + A + Z) and the values of non-photochemical quenching were lower in transgenic plants than in WT plants under high light and chilling stress with low irradiance. The O₂ evolution rate and the maximal photochemical efficiency of PSII (Fv/Fm) in transgenic plants decreased more quickly during both stresses and recovered slower than that in WT under optimal conditions. These results suggested that overexpression of LeZE impaired the function of the xanthophyll cycle and aggravated PSII photoinhibition in tomato under high light and chilling stress.     

Outgrowing endothelial progenitor-derived cells display high sensitivity to angiogenesis modulators and delayed senescence

Outgrowing endothelial progenitor-derived cells (EPDCs) originate from a novel hierarchy of endothelial progenitor cells. In this study, EPDCs isolated from human cord blood were examined for phenotype and functional features upon aging. Young or aged EPDCs were similar to human umbilical vein endothelial cells (HUVECs), in exhibiting typical endothelial phenotypes. However, EPDCs were more sensitive to angiogenesis inducers or inhibitors in proliferation and migration. In addition, EPDCs underwent senescence markedly slowly with sustained endothelial NO synthase expression and activation, and their ability to undergo capillary morphogenesis was retained throughout longterm culture. Thus, these results suggest that a homogenous population of EPDCs derived from clonogenic expansion may provide an effective vasculogenesis tool.     

Senescing leaves possess potential for stress adaptation: the developing leaves acclimated to high light exhibit increased tolerance to osmotic stress during senescence

Plants may experience environmental stress factors operating in nature either simultaneously or in sequence. In the study, we have acclimated the developing primary leaves of wheat seedlings to high light stress and examined their photosynthetic response to polyethylene glycol (PEG) mediated osmotic stress during different developmental phases including senescence. The high light acclimated leaves show higher level of total carotenoids as compared to their non-acclimated counterparts experiencing osmotic stress during senescence. They also exhibit greater membrane stability as indicated by the measurements of fluorescence polarisation and energy transfer efficiency in photosystem I (PSI) and Photosystem II (PSII). From the data of DCPIP photoreduction and pulse amplitude modulated (PAM) fluorimetry, a similar trend is observed for PSII photochemistry of the leaves experiencing osmotic stress during senescence. Our results may suggest that the stress adaptive potential induced by one stress during development is retained by the leaves and helps to mitigate another stress effect operating in sequence during another developmental phase, namely senescence.