Losing the warning signal: Drought compromises the cross-talk of signaling molecules in quercus ilex exposed to ozone

Understanding the interactions between drought and acute ozone (O3) stress in terms of signaling molecules and cell death would improve the predictions of plant responses to climate change. The aim was to investigate whether drought stress influences the responses of plants to acute episodes of O3 exposure. In this study, the behavior of 84 Mediterranean evergreen Quercus ilex plants was evaluated in terms of crosstalk responses among signaling molecules. Half of the sample was subjected to drought (20% of the effective daily evapotranspiration, for 15 days) and was later exposed to an acute O3 exposure (200 nL L-1 for 5 h). First, our results indicate that in well-water conditions, O3 induced a signaling pathway specific to O3-sensitive behavior. Second, different trends and consequently different roles of phytohormones and signaling molecules (ethylene, ET; abscisic acid, ABA; salycilic acid, SA and jasmonic acid, JA) were observed in relation to water stress and O3. A spatial and functional correlation between these signaling molecules was observed in modulating O3-induced responses in well-watered plants. In contrast, in drought-stressed plants, these compounds were not involved either in O3-induced signaling mechanisms or in leaf senescence (a response observed in water-stressed plants before the O3-exposure). Third, these differences were ascribable to the fact that in drought conditions, most defense processes induced by O3 were compromised and/or altered. Our results highlight how Q. ilex plants suffering from water deprivation respond differently to an acute O3 episode compared to well-watered plants, and suggest new effect to be considered in plant responses to environmental changes. This poses the serious question as to whether or not multiple high-magnitude O3 events (as predicted) can change these cross-talk responses, thus opening it up possible further investigations

Protein synthesis inhibition and loss of homeostatic functions in astrocytes from an Alzheimer's disease mouse model: a role for ER-mitochondria interaction

Deregulation of protein synthesis and ER stress/unfolded protein response (ER stress/UPR) have been reported in astrocytes. However, the relationships between protein synthesis deregulation and ER stress/UPR, as well as their role in the altered homeostatic support of Alzheimer's disease (AD) astrocytes remain poorly understood. Previously, we reported that in astrocytic cell lines from 3xTg-AD mice (3Tg-iAstro) protein synthesis was impaired and ER-mitochondria distance was reduced. Here we show that impaired protein synthesis in 3Tg-iAstro is associated with an increase of p-eIF2α and downregulation of GADD34. Although mRNA levels of ER stress/UPR markers were increased two-three-fold, we found neither activation of PERK nor downstream induction of ATF4 protein. Strikingly, the overexpression of a synthetic ER-mitochondrial linker (EML) resulted in a reduced protein synthesis and augmented p-eIF2α without any effect on ER stress/UPR marker genes. In vivo, in hippocampi of 3xTg-AD mice, reduced protein synthesis, increased p-eIF2α and downregulated GADD34 protein were found, while no increase of p-PERK or ATF4 proteins was observed, suggesting that in AD astrocytes, both in vitro and in vivo, phosphorylation of eIF2α and impairment of protein synthesis are PERK-independent. Next, we investigated the ability of 3xTg-AD astrocytes to support metabolism and function of other cells of the central nervous system. Astrocyte-conditioned medium (ACM) from 3Tg-iAstro cells significantly reduced protein synthesis rate in primary hippocampal neurons. When added as a part of pericyte/endothelial cell (EC)/astrocyte 3D co-culture, 3Tg-iAstro, but not WT-iAstro, severely impaired formation and ramification of tubules, the effect, replicated by EML overexpression in WT-iAstro cells. Finally, a chemical chaperone 4-phenylbutyric acid (4-PBA) rescued protein synthesis, p-eIF2α levels in 3Tg-iAstro cells and tubulogenesis in pericyte/EC/3Tg-iAstro co-culture. Collectively, our results suggest that a PERK-independent, p-eIF2α-associated impairment of protein synthesis compromises astrocytic homeostatic functions, and this may be caused by the altered ER-mitochondria interaction

Reactive oxygen species and antioxidant machinery in Liriodendrum tulipifera plants exposed to ozone

Trees are essential in the urban environment not only because of their aesthetic and social values, but also for their effects on air quality. Data of the present work show some of the integrated mechanisms that may confer sensitivity/tolerance in Liriodendron tulipifera (known as the tulip tree) saplings exposed to ozone (O3) (120 ppb, 5 h day-1 for 45 consecutive days) in order to improve the management of green spaces responding to oxidative stress. At the end of fumigation, visible injury due to O3 was observed: symptoms were minute (Ø 1-2 mm) roundish dark-blackish necrosis, mainly located in the interveinal areas of recently fully expanded leaves. The injured leaf area was about 40% of the total surface. Reactive oxygen species levels, membrane injury, enzymes/metabolites linked to ascorbate-glutathione (AsA-GSH) cycle and to the synthesis of phenylpropanoids show modifications caused by O3 in terms of: (i) increase in the superoxide radical production (+41%, in comparison to air filtered controls); (ii) reduction in accumulation of hydrogen peroxide (-55%) and of the enzymes involved in its metabolism [as confirmed by the decrease in superoxide dismutase and catalase activities (-33 and -82%, respectively)]; (iii) increase of antioxidant capacity [as documented by the improved 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) scavenging activity]; (iv) alteration in the solute reactions of the membrane cells [as showed by the production of thiobarbituric acid reactive substances (+34%)]; (v) inhibition of enzymes linked to AsA-GSH cycle [as confirmed by the decrease of ascorbate peroxidase (48%), monodehydroascorbate, dehydroascorbate and glutathione reductase activities (-44, -56 and -80%, respectively)]; (vi) changes of the normal reduced state of cells [as evidenced by the decrease in the reduced ascorbate/dehydroascorbate and in the reduced/oxidized glutathione ratios (-37 and -60%, respectively)]; (vii) activation of enzymes involved in phenolic metabolism [as highlighted by the increase of phenylalanine ammonia-lyase and cinnamyl alcohol dehydrogenase activities (+75 and +67%, respectively)]; (viii) development of cellular barriers with a higher degree of polymerization of monolignols [as indicated by the increase of lignin (+47%)] and (ix) accumulation of phenolic and anthocyanin compounds (+68 and +43%, respectively). These results indicate that O3 exposure modifies reactive oxygen species metabolism rates, but the enzymatic and non-enzymatic antioxidant systems (that work in co-operation) could not provide a defence against free radicals, not preventing the oxidative damage. In conclusion, under these circumstances, L. tulipifera should be regarded as sensitive to this pollutant

Root induction by Agrobacterium rhizogenes in walnut

Agrobacterium rhizogenes (wild-type, strain 1855), when applied to the basal part of microcuttings of walnut (J. regia L.), produced numerous adventitious roots in vitro: 58.6% of rooting was induced in microcuttings in hormone free medium and 62.9% and abundant callus formation in the presence of IBA. A, rhizogenes did not induce rooting when IAA was present in the rooting medium. No explants were induced to root by the treatments with IBA, IAA or in hormone free medium without the presence of A. rhizogenes. Root vascular elements were connected to the microcutting vascular system in the treatment with A. rhizogenes in HFM and were successfully transferred to soil: However, microcuttings treated with A. rhizogenes + IBA showed vascular connections not correctly formed. Southern blots performed using the fragment containing rol genes, as probe, coming from pBIN19::Eco15, provided molecular evidence of the transgenic nature of the roots induced by A. rhizogenes

Can Ozone Alter the Terpenoid Composition and Membrane Integrity of in vitro Melissa officinalis Shoots?

Ozone affects volatile organic compounds that protect plants from biotic and abiotic stress. In vitro Melissa officinalis shoots were exposed to ozone (200 ppb, 3 h) in controlled environmental conditions: leaf pigments, membrane integrity and headspace composition were assayed during fumigation and after the recovery period (3 h from the beginning of the exposure, FBE). At the end of the exposure, no injury was observed in untreated and treated shoots, although an evident increase in lipid peroxidation was reported (+38.5 and +37.2% of TBARS levels in comparison with controls, respectively after 1 and 3 h FBE). The levels of total carotenoids significantly rose as a normal response mechanism to oxidative stress. SPME-GS-MS analysis showed that, as a consequence of the fumigation, the trends in non-terpenoid compounds increased after 1 and 3 h FBE. This suggests that the concentration and the duration of the treatment were enough to cause a breakdown of cells (as evidenced by increased TBARS levels) and involves an association between volatile products of the lipoxygenase pathway (LOX products) and membrane degradation

Accumulation of rosmarinic acid and behaviour of ROS processing systems in Melissa officinalis L. under heat stress

Heat stress (HS) due to increased air temperature is a major agricultural problem. On the other hand, short-term HS can represent a natural easy-to-use elicitor of bioactive compounds in plants. Similar elicitations can be induced by biotechnological approaches such as hydroponic cultures. The present study pioneering investigated the capability of using a short-term HS (38 °C, 5 h) as a tool to rapidly elicit rosmarinic acid (RA) content in leaves of Melissa officinalis L. (a species for which RA is the dominant active phenolic compound) hydroponic cultures, highlighting the cross-talk among antioxidant and signalling molecules involved in the heat acclimation. During HS treatment, we found an elicitation of RA biosynthesis associated with (i) an imbalance in reactive oxygen species (ROS) production and scavenging, (ii) an involvement of reduced ascorbate (AsA) in maintaining a high normal reduced state of cells, (iii) an induction of heat shock proteins (i.e. HSP101-like), and (iv) a stimulation of phytohormones. The RA biosynthesis lasted also during the recovery, although plants activated cellular processes to partially control ROS production, as confirmed by the increased activity of AsA regenerating enzymes, the accumulation of total carotenoids and the stimulation of total antioxidant capacity. The unchanged values of abscisic acid, ethylene and salicylic and jasmonic acids during the recovery phase also documented a reduced demand for protection. The present study represents a wide-ranging investigation of the potential use of HS (without drought interaction) as a technological application for improving bioactive compound production

Ozone as eustress for enhancing secondary metabolites and bioactive properties in Salvia officinalis

Eliciting plants consists in the application of chemical, physical, and biological factors that induce stressful conditions, and so trigger defense mechanisms and the production of bioactive compounds and phytochemicals. In this study, the phenolic and volatile organic compound (VOCs) profiles of sage leaves under a chronic ozone (O3) exposure (120 ppb 5 h day− 1 for 36 consecutive days) were investigated, elucidating also their antioxidant activity. The composition, yield, and antioxidant capacity of essential oils (EOs) obtained from sage leaves were also evaluated. Ozone exposure resulted in an overall increase of phenolic compounds. In terms of VOCs, O3 mainly reduced monoterpene emissions, while increased the production of sesquiterpenes (in both leaves and EOs). These O3-induced accumulations were triggered during the first weeks of exposure, whereas they dis- appeared at the last time of analysis, suggesting that sage plants lost their ability (or interest) in activating secondary metabolism under high doses of O3. The antioxidant capacity of all tested extracts resulted increased by O3 exposure. These outcomes support our speculation that the application of O3 for a limited period (i.e., a maximum of four weeks, at the investigated concentration) may become an effective biotechnological tool to improve the quality of sage leaf extracts

When "thirsty" means "less able to activate the signalling wave trigged by a pulse of ozone": A case of study in two Mediterranean deciduous oak species with different drought sensitivity

There is a lack of knowledge about the possibility that plants facing abiotic stressors, such as drought, have an altered perception of a pulse of O3 and incur in alterations of their signalling network. This poses some concerns as to whether defensive strategy to cope episodic O3 peaks in healthy plants may fail under stress. In this study, a set of saplings of two Mediterranean deciduous species, Quercus cerris and Q. pubescens, was subjected to water withholding (20% of daily evapotranspiration for 15 days) while another set was kept well-watered. Saplings were then subjected to a pulse of O3 (200 nl l-1 for 5 h) or maintained in filtered air. Q. pubescens had a more severe decline of photosynthesis and leaf PDΨw (about -65% and 5-fold lower than in well-watered ones) and events of cell death were observed under drought when compared to Q. cerris, which is supportive for a higher sensitivity to drought exhibited by this species. When O3 was applied after drought, patterns of signalling compounds were altered in both species. Only in Q. pubescens, the typical O3-induced accumulation of apoplastic reactive oxygen species, which is the first necessary step for the activation of signalling cascade, was completely lost. In Q. cerris the most frequent changes encompassed the weakening of peaks of key signalling molecules (ethylene and salicylic acid), whereas in Q. pubescens both delayed (salicylic and jasmonic acid) or weakened (ethylene and salicylic acid) peaks were observed. This is translated to a higher ability of Q. cerris to maintain a prompt activation of defensive reaction to counteract oxidative damage due to the pollutant. Our results reveal the complexity of the signalling network in plants facing multiple stresses and highlight the need to further investigate possible alteration of defensive mechanism of tree species to predict their behavior