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Climate warming is altering snowpack permanence in alpine tundra, modifying shrub growth and distribution. Plant acclimation to snowpack changes depends on the capability to guarantee growth and carbon storage, suggesting that the content of non-structural carbohydrates (NSC) in plant organs can be a key trait to depict the plant response under different snow regimes. To test this hypothesis, we designed a 3-years long manipulative experiment aimed at evaluating the effect of snow melt timing (i.e., early, control, and late) on NSC content in needles, bark and wood of Juniperus communis L. growing at high elevation in the Alps. Starch evidenced a general decrease from late spring to summer in control and early melting, while starch was low but stable in plants subjected to a late snow melt. Leaves, bark and wood have different level of soluble NSC changing during growing season: in bark, sugars content decreased significantly in late summer, while there was no seasonal effect in needles and wood. Soluble NSC and starch were differently related with the plant growth, when considering different tissues and snow treatment. In leaf and bark we observed a starch depletion in control and early melting plants, consistently to a higher growth (i.e., twig elongation), while in late snow melt, we did not find any significant relationship between growth and NSC concentration. Our findings confirmed that snowpack duration affects the onset of the growing season promoting a change in carbon allocation in plant organs and, between bark and wood in twigs. Finally, our results suggest that plants, at this elevation, could take advantage from an early snow melt caused by climate warming, most likely due to photosynthetic activity by maintaining the level of reserves and enhancing the carbon investment for growth., Competing Interests: Declaration of competing interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024. Published by Elsevier B.V.)

Woody plants display some photosynthetic activity in stems, but the biological role of stem photosynthesis and the specific contributions of bark and wood to carbon uptake and oxygen evolution remain poorly understood. We aimed to elucidate the functional characteristics of chloroplasts in stems of different ages in Fraxinus ornus. Our investigation employed diverse experimental approaches, including microsensor technology to assess oxygen production rates in whole stem, bark, and wood separately. Additionally, we utilized fluorescence lifetime imaging microscopy (FLIM) to characterize the relative abundance of photosystems I and II (PSI : PSII chlorophyll ratio) in bark and wood. Our findings revealed light-induced increases in O 2 production in whole stem, bark, and wood. We present the radial profile of O 2 production in F. ornus stems, demonstrating the capability of stem chloroplasts to perform light-dependent electron transport. Younger stems exhibited higher light-induced O 2 production and dark respiration rates than older ones. While bark emerged as the primary contributor to net O 2 production under light conditions, our data underscored that wood chloroplasts are also photosynthetically active. The FLIM analysis unveiled a lower PSI abundance in wood than in bark, suggesting stem chloroplasts are not only active but also acclimate to the spectral composition of light reaching inner compartments., (© 2024 The Authors. New Phytologist © 2024 New Phytologist Foundation.)

Plant roots are exposed to hypoxia in waterlogged soils, and they are further challenged by specific phytotoxins produced by microorganisms in such conditions. One such toxin is hexanoic acid (HxA), which, at toxic levels, causes a strong decline in root O 2 consumption. However, the mechanism underlying this process is still unknown. We treated pea (Pisum sativum L.) roots with 20 mM HxA at pH 5.0 and 6.0 for a short time (1 h) and measured leakage of key electrolytes such as metal cations, malate, citrate and nonstructural carbohydrates (NSC). After treatment, mitochondria were isolated to assess their functionality evaluated as electrical potential and O 2 consumption rate. HxA treatment resulted in root tissue extrusion of K + , malate, citrate and NSC, but only the leakage of the organic acids and NSC increased at pH 5.0, concomitantly with the inhibition of O 2 consumption. The activity of mitochondria isolated from treated roots was almost unaffected, showing just a slight decrease in oxygen consumption after treatment at pH 5.0. Similar results were obtained by treating the pea roots with another organic acid with a short carbon chain, that is, butyric acid. Based on these results, we propose a model in which HxA, in its undissociated form prevalent at acidic pH, stimulates the efflux of citrate, malate and NSC, which would, in turn, cause starvation of mitochondrial respiratory substrates of the Krebs cycle and a consequent decline in O 2 consumption. Cation extrusion would be a compensatory mechanism in order to restore plasma membrane potential., (© 2023 The Authors. Physiologia Plantarum published by John Wiley & Sons Ltd on behalf of Scandinavian Plant Physiology Society.)

Xylem embolism is one of the possible outcomes of decreasing xylem pressure when plants face drought. Recent studies have proposed a role for non-structural carbohydrates (NSCs) in osmotic pressure generation, required for refilling embolized conduits. Potted cuttings of grapevine Grenache and Barbera, selected for their adaptation to different climatic conditions, were subjected to a drought stress followed by re-irrigation. Stem embolism rate and its recovery were monitored in vivo by X-ray micro-computed tomography (micro-CT). The same plants were further analyzed for xylem conduit dimension and NSC content. Both cultivars significantly decreased Ψpd in response to drought and recovered from xylem embolism after re-irrigation. However, although the mean vessel diameter was similar between the cultivars, Barbera was more prone to embolism. Surprisingly, vessel diameter was apparently reduced during recovery in this cultivar. Hydraulic recovery was linked to sugar content in both cultivars, showing a positive relationship between soluble NSCs and the degree of xylem embolism. However, when starch and sucrose concentrations were considered separately, the relationships showed cultivar-specific and contrasting trends. We showed that the two cultivars adopted different NSC-use strategies in response to drought, suggesting two possible scenarios driving conduit refilling. In Grenache, sucrose accumulation seems to be directly linked to embolism formation and possibly sustains refilling. In Barbera, maltose/maltodextrins could be involved in a conduit recovery strategy via the formation of cell-wall hydrogels, likely responsible for the reduction of conduit lumen detected by micro-CT., (© The Author(s) 2023. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The stress gradient hypothesis (SGH) states that plant-plant interactions shift from competition to facilitation in increasing stress conditions. In salt marshes, edaphic properties can weaken the application of the SGH by amplifying the intensity of flooding and controlling plant zonation. We identified facilitative and competitive interactions along flooding gradients and tested the role of edaphic properties in exacerbating stress and shaping plant-plant interactions. Morphological traits of two target halophytes ( Limonium narbonense and Sarcocornia fruticosa ), flooding intensity, soil texture and soil organic C were recorded. The relative plant fitness index was assessed for the two species based on the relative growth in plurispecific rather than monospecific plant communities. Plant fitness increased with increasing stress supporting the SGH. L. narbonense showed larger fitness in plurispecific stands whereas S. fruticosa performed better in conspecific stands. Significant intra- or interspecific interactions were observed along the stress gradient defined by the combination of flooding and clay content in soil. When considering the limited soil organic C as stressor, soil properties were more important than flooding in defining plant-plant interactions. We highlight the need for future improvements of the SGH approach by including edaphic stressors in the model and their possible interactions with the main abiotic drivers of zonation.

With the recent climate warming, tundra ecotones are facing a progressive acceleration of spring snowpack melting and extension of the growing season, with evident consequences to vegetation. Along with summer temperature, winter precipitation has been recently recognised as a crucial factor for tundra shrub growth and physiology. However, gaps of knowledge still exist on long-living plant responses to different snowpack duration, especially on how intra-specific and year-to-year variability together with multiple functional trait adjustments could influence the long-term responses. To fill this gap, we conducted a 3 years snow manipulation experiment above the Alpine treeline on the typical tundra species Juniperus communis , the conifer with the widest distributional range in the north emisphere. We tested shoot elongation, leaf area, stomatal density, leaf dry weight and leaf non-structural carbohydrate content of plants subjected to anticipated, natural and postponed snowpack duration. Anticipated snowpack melting enhanced new shoot elongation and increased stomatal density. However, plants under prolonged snow cover seemed to compensate for the shorter growing period, likely increasing carbon allocation to growth. In fact, these latter showed larger needles and low starch content at the beginning of the growing season. Variability between treatments slightly decreased over time, suggesting a progressive acclimation of juniper to new conditions. In the context of future warming scenarios, our results support the hypothesis of shrub biomass increase within the tundra biome. Yet, the picture is still far from being complete and further research should focus on transient and fading effects of changing conditions in the long term., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Unterholzner, Prendin, Dibona, Menardi, Casolo, Gargiulo, Boscutti and Carrer.)

Nonstructural carbohydrates (NSCs) have been suggested to affect xylem transport under fluctuating water availability, but conclusive evidence is still lacking. We tested the effect of shade-induced NSC depletion on xylem vulnerability to embolism and hydraulic recovery on Populus nigra saplings. Vulnerability was assessed in light-exposed (L) and shaded (S) plants with the hydraulic method, and in vivo with the optical method and X-ray micro-computed tomography. Plants were stressed to 80% loss of hydraulic conductance (PLC) and re-irrigated to check for possible recovery. We measured PLC, bark and wood NSC content, as well as xylem sap pH, surface tension (γ sap ) and sugar concentration, before, during and after drought. Shading induced depletion of stem NSC (mainly starch) reserves. All methods converged in indicating higher xylem vulnerability in S than in L plants. This difference was not explained by xylem vessel and pit anatomy or by γ sap . Shading impeded sap acidification and sugar accumulation during drought in S plants and prevented hydraulic recovery, which was observed in L plants. Our results highlight the importance of stem NSCs to sustain xylem hydraulic functioning during drought and suggest that light and/or adequate stem NSC thresholds are required to trigger xylem sap chemical changes involved in embolism recovery., (© 2021 The Authors New Phytologist © 2021 New Phytologist Foundation.)

Premise: Despite great attention given to the relationship between plant growth and carbon balance in alpine tree species, little is known about shrubs at the treeline. We hypothesized that the pattern of main nonstructural carbohydrates (NSCs) across elevations depends on the interplay between phenotypic trait plasticity, plant-plant interaction, and elevation., Methods: We studied the pattern of NSCs (i.e., glucose, fructose, sucrose, and starch) in alpine stands of Vaccinium myrtillus (above treeline) across an elevational gradient. In the same plots, we measured key growth traits (i.e., anatomical stem features) and shrub cover, evaluating putative relationships with NSCs., Results: Glucose content was positively related with altitude, but negatively related with shrub cover. Sucrose decreased at high altitude and in older populations and increased with higher percentage of vascular tissue. Starch content increased at middle and high elevations and in stands with high shrub cover. Moreover, starch content was negatively related with the number of xylem rings and the percentage of phloem tissue, but positively correlated with the percentage of xylem tissue., Conclusions: We found that the increase in carbon reserves across elevations was uncoupled from plant growth, supporting the growth limitation hypothesis, which postulates NSCs accumulate at high elevation as a consequence of low temperature. Moreover, the response of NSC content to the environmental stress caused by elevation was buffered by phenotypic plasticity of plant traits, suggesting that, under climate warming conditions, shrub expansion due to enhanced plant growth would be pronounced in old but sparse stands., (© The Authors. American Journal of Botany published by Wiley Periodicals, Inc. on behalf of Botanical Society of America.)

In grapevine, the anatomy of xylem conduits and the non-structural carbohydrates (NSCs) content of the associated living parenchyma are expected to influence water transport under water limitation. In fact, both NSC and xylem features play a role in plant recovery from drought stress. We evaluated these traits in petioles of Cabernet Sauvignon (CS) and Syrah (SY) cultivars during water stress (WS) and recovery. In CS, the stress response was associated to NSC consumption, supporting the hypothesis that starch mobilization is related to an increased supply of maltose and sucrose, putatively involved in drought stress responses at the xylem level. In contrast, in SY, the WS-induced increase in the latter soluble NSCs was maintained even 2 days after re-watering, suggesting a different pattern of utilization of NSC resources. Interestingly, the anatomical analysis revealed that conduits are constitutively wider in SY in well-watered (WW) plants, and that water stress led to the production of narrower conduits only in this cultivar.

The xylem is a complex system that includes a network of dead conduits ensuring long-distance water transport in plants. Under ongoing climate changes, xylem embolism is a major and recurrent cause of drought-induced tree mortality. Non-structural carbohydrates (NSC) play key roles in plant responses to drought and frost stress, and several studies putatively suggest their involvement in the regulation of xylem water transport. However, a clear picture on the roles of NSCs in plant hydraulics has not been drawn to date. We summarize the current knowledge on the involvement of NSCs during embolism formation and subsequent hydraulic recovery. Under drought, sugars are generally accumulated in xylem parenchyma and in xylem sap. At drought-relief, xylem functionality is putatively restored in an osmotically driven process involving wood parenchyma, xylem sap and phloem compartments. By analyzing the published data on stem hydraulics and NSC contents under drought/frost stress and subsequent stress relief, we found that embolism build-up positively correlated to stem NSC depletion, and that the magnitude of post-stress hydraulic recovery positively correlated to consumption of soluble sugars. These findings suggest a close relationship between hydraulics and carbohydrate dynamics. We call for more experiments on hydraulic and NSC dynamics in controlled and field conditions., Competing Interests: The authors declare no conflict of interest

The maintenance of hydraulic function during and after a drought event is crucial for tree survival, but the importance of non-structural carbohydrates (NSCs) in the recovery phase is still debated. We tested whether higher NSC availability facilitates post-drought hydraulic recovery, by applying a short-term drought (S dr ) and a long-term drought combined with shading (L dr+sh ) in Fraxinus ornus and Ostrya carpinifolia. Plants were then re-irrigated and recovery was checked 24 h later, by measuring water potential, stem percentage loss of hydraulic conductance (PLC) and NSC content. The relative magnitude of hydraulic and carbon constraints was also assessed in desiccated plants. During drought, PLC increased only in F. ornus, while it was maintained almost constant in O. carpinifolia due to tighter stomatal control of xylem pressure (i.e. more isohydric). In F. ornus, only S dr plants maintained high NSC contents at the end of drought and, when re-irrigated, recovered PLC to control values. Whereas hydraulic failure was ubiquitous, only F. ornus depleted NSC reserves at mortality. Our results suggest that preserving higher NSC content at the end of a drought can be important for the hydraulic resilience of trees., (Copyright © 2019 Elsevier Masson SAS. All rights reserved.)

Embolism repair ability has been documented in numerous species. Although the actual mechanism driving this phenomenon is still debated, experimental findings suggest that non-structural carbohydrates (NSC) stored in wood parenchyma would provide the osmotic forces to drive the refilling of embolized conduits. We selected 12 broadleaved species differing in vulnerability to xylem embolism (P 50 ) and amount of wood parenchyma in order to check direct evidence about the possible link(s) between parenchyma cells abundance, NSC availability and species-specific capacity to reverse xylem embolism. Branches were dehydrated until ∼50% loss of hydraulic conductivity was recorded (PLC ∼50%). Hydraulic recovery (ΔPLC) and NSC content was, then, assessed after 1h of rehydration. Species showed a different ability to recover their hydraulic conductivity from PLC∼50%. Removing the bark in the species showing hydraulic recovery inhibited the embolism reversal. Strong correlations between the ΔPLC and: a) the amount of parenchyma cells (mainly driven by the pith area), b) the consumption of soluble NSC have been recorded. Our results support the hypothesis that refilling of embolized vessels is mediated by the mobilization of soluble NSC and it is mainly recorded in species with a higher percentage of parenchyma cells that may be important in the hydraulic recovery mechanism as a source of carbohydrates and/or as a source of water., (Copyright © 2019 Elsevier Masson SAS. All rights reserved.)

Some plant species are capable of significant reduction of xylem embolism during recovery from drought despite stem water potential remains negative. However, the functional biology underlying this process is elusive. We subjected poplar trees to drought stress followed by a period of recovery. Water potential, hydraulic conductivity, gas exchange, xylem sap pH, and carbohydrate content in sap and woody stems were monitored in combination with an analysis of carbohydrate metabolism, enzyme activity, and expression of genes involved in sugar metabolic and transport pathways. Drought resulted in an alteration of differential partitioning between starch and soluble sugars. Upon stress, an increase in the starch degradation rate and the overexpression of sugar symporter genes promoted the efflux of disaccharides (mostly maltose and sucrose) to the apoplast. In turn, the efflux activity of the sugar-proton cotransporters caused a drop in xylem pH. The newly acidic environment induced the activity of apoplastic invertases leading to the accumulation of monosaccharides in the apoplast, thus providing the main osmoticum necessary for recovery. During drought and recovery, a complex network of coordinated molecular and biochemical signals was activated at the interface between xylem and parenchyma cells that appeared to prime the xylem for hydraulic recovery., (© 2019 John Wiley & Sons Ltd.)

Ongoing climate change is apparently increasing tree mortality rates, and understanding mechanisms of drought-induced tree decline can improve mortality projections. Differential drought impact on conspecific individuals within a population has been reported, but no clear mechanistic explanation for this pattern has emerged. Following a severe drought (summer 2012), we monitored over a 3-year period healthy (H) and declining (D) Pinus nigra trees co-occurring in a karstic woodland to highlight eventual individual-specific physiological differences underlying differential canopy dieback. We investigated differences in water and carbon metabolism, and xylem anatomy as a function of crown health status, as well as eventual genotypic basis of contrasting drought responses. H and D trees exploited the same water pools and relied on similar hydraulic strategies to cope with drought stress. Genetic analyses did not highlight differences between groups in terms of geographical provenance. Hydraulic and anatomical analyses showed conflicting results. The hydraulic tracheid diameter and theoretical hydraulic conductivity were similar, but D trees were characterized by lower water transport efficiency, greater vulnerability to xylem conduit implosion and reduced carbohydrate stores. Our results suggest that extreme drought events can have different impacts on conspecific individuals, with differential vulnerability to xylem embolism likely playing a major role in setting the fate of trees under climate change.

In striking analogy with Saccharomyces cerevisiae , etiolated pea stem mitochondria did not show appreciable Ca 2+ uptake. Only treatment with the ionophore ETH129 (which allows electrophoretic Ca 2+ equilibration) caused Ca 2+ uptake followed by increased inner membrane permeability, membrane depolarization and Ca 2+ release. Like the permeability transition (PT) of mammals, yeast and Drosophila, the PT of pea stem mitochondria was stimulated by diamide and phenylarsine oxide and inhibited by Mg-ADP and Mg-ATP, suggesting a common underlying mechanism; yet, the plant PT also displayed distinctive features: (i) as in mammals it was desensitized by cyclosporin A, which does not affect the PT of yeast and Drosophila; (ii) similarly to S. cerevisiae and Drosophila it was inhibited by Pi, which stimulates the PT of mammals; (iii) like in mammals and Drosophila it was sensitized by benzodiazepine 423, which is ineffective in S. cerevisiae ; (iv) like what observed in Drosophila it did not mediate swelling and cytochrome c release, which is instead seen in mammals and S. cerevisiae . We find that cyclophilin D, the mitochondrial receptor for cyclosporin A, is present in pea stem mitochondria. These results indicate that the plant PT has unique features and suggest that, as in Drosophila, it may provide pea stem mitochondria with a Ca 2+ release channel.

Enhanced shrub growth and expansion are widespread responses to climate warming in many arctic and alpine ecosystems. Warmer temperatures and shrub expansion could cause major changes in plant community structure, affecting both species composition and diversity. To improve our understanding of the ongoing changes in plant communities in alpine tundra, we studied interrelations among climate, shrub growth, shrub cover and plant diversity, using an elevation gradient as a proxy for climate conditions. Specifically, we analyzed growth of bilberry (Vaccinium myrtillus L.) and its associated plant communities along an elevation gradient of ca. 600 vertical meters in the eastern European Alps. We assessed the ramet age, ring width and shoot length of V. myrtillus, and the shrub cover and plant diversity of the community. At higher elevation, ramets of V. myrtillus were younger, with shorter shoots and narrower growth rings. Shoot length was positively related to shrub cover, but shrub cover did not show a direct relationship with elevation. A greater shrub cover had a negative effect on species richness, also affecting species composition (beta-diversity), but these variables were not influenced by elevation. Our findings suggest that changes in plant diversity are driven directly by shrub cover and only indirectly by climate, here represented by changes in elevation.

Drought-induced tree decline is a complex event, and recent hypotheses suggest that hydraulic failure and carbon starvation are co-responsible for this process. We tested the possible role of non-structural carbohydrates (NSC) content on post-drought hydraulic recovery, to verify the hypothesis that embolism reversal represents a mechanistic link between carbon starvation and stem hydraulics. Measurements were performed in laurel plants subjected to similar water stress levels either over short or long term, to induce comparable embolism levels. Plants subjected to mild and prolonged water shortage (S) showed reduced growth, adjustment of turgor loss point driven by changes in both osmotic potential at full turgor and bulk modulus of elasticity, a lower content of soluble NSC and a higher content of starch with respect to control (C) plants. Moreover, S plants showed a lower ability to recover from xylem embolism than C plants, even after irrigation. Our data suggest that plant carbon status might indirectly influence plant performance during and after drought via effects on xylem hydraulic functioning, supporting the view of a possible mechanistic link between the two processes., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

Terrestrial saltmarsh plants inhabiting flood-prone habitats undergo recurrent and prolonged flooding driven by tidal regimes. In this study, the role of internal plant aeration in contrasting hypoxic/anoxic conditions during submergence was investigated in the two halophytes Limonium narbonense Mill. and Sarcocornia fruticosa (L.) A.J. Scott. Monitoring of tissue O2 dynamics was performed in shoots and roots using microelectrodes under drained conditions, waterlogging, partial and complete submergence, in light or darkness. For both species, submergence in darkness resulted in significant declines in tissue O2 status and when in light, in rapid O2 increases first in shoot tissues and subsequently in roots. During partial submergence, S. fruticosa benefitted from snorkelling and efficiently transported O2 to roots, whereas the O2 concentration in roots of L. narbonense declined by more than 90%. Significantly thinner leaves and articles were recorded under high degree of flooding stress and both species showed considerably high tissue porosity. The presence of aerenchyma seemed to support internal aeration in S. fruticosa whereas O2 diffusion in L. narbonense seemed impeded, despite the higher porosity (up to 50%). Thus, the results obtained for L. narbonense, being well adapted to flooding, suggests that processes other than internal aeration could be involved in better flooding tolerance e.g. fermentative processes, and that traits resulting in flooding tolerance in plants are not yet fully understood.

Background: Senescence is a key developmental process occurring during the life cycle of plants that can be induced also by environmental conditions, such as starvation and/or darkness. During senescence, strict control of genes regulates ordered degradation and dismantling events, the most remarkable of which are genetically programmed cell death (PCD) and, in most cases, an upregulation of flavonoid biosynthesis in the presence of light. Flavonoids are secondary metabolites that play multiple essential roles in development, reproduction and defence of plants, partly due to their well-known antioxidant properties, which could affect also the same cell death machinery. To understand further the effect of endogenously-produced flavonoids and their interplay with different environment (light or dark) conditions, two portions (red and green) of a senescing grapevine callus were used to obtain suspension cell cultures. Red Suspension cell Cultures (RSC) and Green Suspension cell Cultures (GSC) were finally grown under either dark or light conditions for 6 days., Results: Darkness enhanced cell death (mainly necrosis) in suspension cell culture, when compared to those grown under light condition. Furthermore, RSC with high flavonoid content showed a higher viability compared to GSC and were more protected toward PCD, in accordance to their high content in flavonoids, which might quench ROS, thus limiting the relative signalling cascade. Conversely, PCD was mainly occurring in GSC and further increased by light, as it was shown by cytochrome c release and TUNEL assays., Conclusions: Endogenous flavonoids were shown to be good candidates for exploiting an efficient protection against oxidative stress and PCD induction. Light seemed to be an important environmental factor able to induce PCD, especially in GSC, which lacking of flavonoids were not capable of preventing oxidative damage and signalling leading to senescence.

Recent reports on tree mortality associated with anomalous drought and heat have raised interest into processes underlying tree resistance/resilience to water stress. Hydraulic failure and carbon starvation have been proposed as main causes of tree decline, with recent theories treating water and carbon metabolism as interconnected processes. We subjected young plants of two native (Quercus pubescens [Qp] and Prunus mahaleb [Pm]) and two invasive (Robinia pseudoacacia [Rp] and Ailanthus altissima [Aa]) woody angiosperms to a prolonged drought leading to stomatal closure and xylem embolism, to induce carbon starvation and hydraulic failure. At the end of the treatment, plants were measured for embolism rates and NSC content, and re-irrigated to monitor recovery of xylem hydraulics. Data highlight different hydraulic strategies in native vs invasive species under water stress, and provide physiological explanations for species-specific impacts of recent severe droughts. Drought-sensitive species (Qp and Rp) suffered high embolism rates and were unable to completely refill xylem conduits upon restoration of water availability. Species that better survived recent droughts were able to limit embolism build-up (Pm) or efficiently restored hydraulic functionality after irrigation (Aa). Species-specific capacity to reverse xylem embolism correlated to stem-level concentration of soluble carbohydrates, but not to starch content., (Copyright © 2016 Elsevier Masson SAS. All rights reserved.)

In 2012, an extreme summer drought induced species-specific die-back in woody species in Northeastern Italy. Quercus pubescens and Ostrya carpinifolia were heavily impacted, while Prunus mahaleb was largely unaffected. By comparing seasonal changes in isotopic composition of xylem sap, rainfall and deep soil samples, we show that P. mahaleb has a deeper root system than the other two species. This morphological trait allowed P  mahaleb to maintain higher water potential (Ψ), gas exchange rates and non-structural carbohydrates content (NSC) throughout the summer, when compared with the other species. More favourable water and carbon states allowed relatively stable maintenance of stem hydraulic conductivity (k) throughout the growing season. In contrast, in Quercus pubescens and Ostrya carpinifolia, decreasing Ψ and NSC were associated with significant hydraulic failure, with spring-to-summer k loss averaging 60%. Our data support the hypothesis that drought-induced tree decline is a complex phenomenon that cannot be modelled on the basis of single predictors of tree status like hydraulic efficiency, vulnerability and carbohydrate content. Our data highlight the role of rooting depth in seasonal progression of water status, gas exchange and NSC, with possible consequences for energy-demanding mechanisms involved in the maintenance of vascular integrity., (© 2015 John Wiley & Sons Ltd.)

The synthesis of ATP in mitochondria is dependent on a low permeability of the inner membrane. Nevertheless, mitochondria can undergo an increased permeability to solutes, named permeability transition (PT) that is mediated by a permeability transition pore (PTP). PTP opening requires matrix Ca(2+) and leads to mitochondrial swelling and release of intramembrane space proteins (e.g., cytochrome c). This feature has been initially observed in mammalian mitochondria and tentatively attributed to some components present either in the outer or inner membrane. Recent works on mammalian mitochondria point to mitochondrial ATP synthase dimers as physical basis for PT, a finding that has been substantiated in yeast and Drosophila mitochondria. In plant mitochondria, swelling and release of proteins have been linked to programmed cell death, but in isolated mitochondria PT has been observed in only a few cases and in plant cell cultures only indirect evidence is available. The possibility that mitochondrial ATP synthase dimers could function as PTP also in plants is discussed here on the basis of the current evidence. Finally, a hypothetical explanation for the origin of PTP is provided in the framework of molecular exaptation.

Spartina patens (Ait.) Muhl. is a grass native to the Atlantic coastal area of North America currently invading salt marsh ecosystems in several regions of Europe. We investigated leaf water relations and hydraulics, gas exchange, nitrogen and starch content in two populations of S. patens growing under contrasting salinity levels in a salt marsh and in a dune system in order to assess its functional plasticity as a factor contributing to its invasive potential. The analysis of leaf water relations revealed a suite of mechanisms adopted by S. patens to overcome salt and drought stress while maintaining relatively invariant leaf morphological traits and plant biomass. In particular, salt marsh plants experiencing severe water stress underwent greater osmoregulation and leaf hydraulic adjustment than dune plants. We also present the first experimental evidence for salt-mediated regulation of xylem hydraulic efficiency in a halophytic grass and suggest that it is an important functional trait allowing plants growing in saline habitats to cope with a restricted water supply. The functional plasticity of leaf water relations and xylem hydraulics emerges as a key trait underlying the competitive ability and invasive potential of S. patens.

Biscutella laevigata is a facultative metallophyte, with populations on non-metalliferous and metalliferous soils. Some of its metallicolous populations have been shown to hyperaccumulate thallium or lead in nature. Only Tl hyperaccumulation has been experimentally confirmed. We aimed to compare the patterns of metal (hyper)accumulation and genetic diversity among populations of B. laevigata subsp. laevigata in NE Italy. None of the populations exhibited foliar hyperaccumulation of Cu, Zn, or Pb. The root-to-shoot accumulation rates for these metals were unchanged or decreased rather than enhanced in the metallicolous populations, in comparison with the non-metallicolous ones. Hyperaccumulation of Tl was confined to the population of the Cave del Predil mine. This population was genetically very distinct from the others, as demonstrated by AFLP-based cluster analysis. The two other mine populations did not surpass the threshold for Tl hyperaccumulation, but showed enhanced foliar Tl concentrations and root-to-shoot translocation rates, in comparison with the non-metallicolous populations. Genetic analysis suggested that adaptation to metalliferous soil must have been independently evolved in the metallicolous populations.

The mitochondrial permeability transition (PT) is a well-recognized phenomenon that allows mitochondria to undergo a sudden increase of permeability to solutes with molecular mass ≤ 1500Da, leading to organelle swelling and structural modifications. The relevance of PT relies on its master role in the manifestation of programmed cell death (PCD). This function is performed by a mega-channel (in some cases inhibited by cyclosporin A) named permeability transition pore (PTP), whose function could derive from the assembly of different mitochondrial proteins. In this paper we examine the distribution and characteristics of PTP in mitochondria of eukaryotic organisms so far investigated in order to draw a hypothesis on the mechanism of its evolution. As a result, we suggest that PTP may have arisen as a new function linked to a multiple molecular exaptation of different mitochondrial proteins, even though they could nevertheless still play their original role. Furthermore, we suggest that the early appearance of PTP could have had a crucial role in the establishment of endosymbiosis in eukaryotic cells, by the coordinated balancing of ATP production by glycolysis (performed by the primary phagocyte) and oxidative phosphorylation (accomplished by the endosymbiont). Indeed, we argue on the possibility that this new energetic equilibrium could have opened the way to the subsequent evolution toward metazoans., (© 2012 Elsevier B.V. All rights reserved.)

Flavonoids are a class of secondary metabolites present in large amounts in grapevine (Vitis vinifera L.), which are involved in several aspects of its physiology (e.g. protection against biotic and abiotic stress). Even if the biosynthetic pathways of flavonoid sub-classes have been largely characterised, the mechanisms of their transport and accumulation to the final target sites are still not completely understood. Unanticipated insights have been obtained by probing plant tissues with pure antibodies targeting bilitranslocase (BTL, TCDB # 2.A.65.1.1), a mammalian transporter involved in the absorption and tissue distribution of dietary flavonoids. The occurrence of a BTL homologue has also been found in grape berries, in both tegumental layers of skin and pulp vascular bundles. In the skin, the expression of this protein starts from véraison (starting of the change in colour and softening of berries) and increases up to a maximum at the harvest stage, matching the same temporal pattern of flavonoid accumulation.

The present work reports changes in bioenergetic parameters and mitochondrial activities during the manifestation of two events of programmed cell death (PCD), linked to Abies alba somatic embryogenesis. PCD, evidenced by in situ nuclear DNA fragmentation (TUNEL assay), DNA laddering and cytochrome c release, was decreased in maturing embryogenic tissue with respect to the proliferation stage. In addition, the major cellular energetic metabolites (ATP, NAD(P)H and glucose-6-phosphate) were highered during maturation. The main mitochondrial activities changed during two developmental stages. Mitochondria, isolated from maturing, with respect to proliferating cell masses, showed an increased activity of the alternative oxidase, external NADH dehydrogenase and fatty-acid mediated uncoupling. Conversely, a significant decrease of the mitochondrial K (ATP)(+) channel activity was observed. These results suggest a correlation between mitochondrial activities and the manifestation of PCD during the development of somatic embryos. In particular, it is suggested that the K (ATP)(+) channel activity could induce an entry of K(+) into the matrix, followed by swelling and a release of cytochrome c during proliferation, whereas the alternative pathways, acting as anti-apoptotic factors, may partially counteract PCD events occurring during maturation of somatic embryos.

This report demonstrates that mitochondria isolated from thermogenic Arum spadices possess an ATP-sensitive potassium channel--responsible for electrical potential (DeltaPsi) collapse and mitochondrial swelling--whose characteristics are similar to those previously described in pea and wheat mitochondria. In order to study the relationship between this K(ATP)(+) channel and the uncoupled respiration, linked to thermogenesis, K(+) transport activities were compared with those of mitochondria that were isolated from pea stems, soybean suspension cell cultures and Arum tubers. The channel from Arum spadices is highly active and its major features are (i) potassium flux is performed primarily in an inward-rectifying manner; (ii) the influx of K(+) is associated with a matrix volume increase in both energized and non-energized mitochondria; and (iii) its activity depends on the redox state of electron transport chain (ETC) and oxygen availability. In particular, this paper shows that the K(ATP)(+) channel is inwardly activated in parallel with the alternative oxidase (AO). The activation is linked to an ETC-oxidized state and to high oxygen consumption. The putative role of this K(ATP)(+) channel is discussed in relation to flowering of thermogenic Arum spadices.

A valuable method to isolate and purify mitochondria from embryonal masses of two coniferous species (Picea abies [L.] Karst. and Abies cephalonica Loud.) is described. Crude mitochondria from both species were shown to be intact, oxygen consuming (with malate plus glutammate, succinate and NADH as substrates) and well coupled (respiratory control ratio ca. 4). The oxidation of the substrates was only partially KCN-insensitive (alternative oxidase) in some cases. However, these fractions were contaminated by membranes (e.g. plasmalemma, tonoplast, Golgi and endoplasmic reticulum). After purification by a discontinuous Percoll gradient (18, 23, 40%, v/v), three mitochondrial populations were separated. The 0/18 interface fraction was composed mainly of broken and uncoupled mitochondria, while the other two (18/23 and 23/40 interface fractions) contained intact and coupled mitochondria, but only 23/40 interface fraction revealed to be better purified starting from both coniferous embryonal masses. In the latter purified fraction, the presence of a cyclosporin A-sensitive K (ATP) (+) channel was demonstrated. These findings were discussed in the light of the potential use of these mitochondrial fractions in bioenergetic studies, or in the involvement of these organelles to stress response in conifers.

Soybean suspension cell cultures were treated by H2O2 or nitric oxide (NO), to assess the mechanism leading to programmed cell death (PCD). Hydrogen peroxide (5 mM) induced PCD. Cells become necrotic at 20 mM H2O2, with cells exhibiting intermediate hallmarks before that (necrapoptotic cells). The level of ATP and of glucose-6-phosphate remained constant in cells undergoing PCD, while it decreased significantly in the necrotic ones. Mitochondria, isolated from 5 mM H2O2-treated (apoptotic) cells, showed that succinate-dependent oxygen consumption was slightly uncoupled, and the electrical potential difference (delta psi) weakly decreased. The addition of KCl to the delta psi formed determined a partial dissipation, which was higher than the dissipation observed in mitochondria from control cells. The addition of cyclosporin A (CsA) to de-energized mitochondria also induced delta psi formation, due to a K+ efflux from the matrix, which was decreased in mitochondria from treated cells. The same pattern of response was also observed in mitochondria isolated from 1 mM sodium nitroprusside (NO)-treated cells, exhibiting apoptotic symptoms. In mitochondria isolated from 20 mM H2O2-treated (necrotic) cells, succinate-dependent oxygen consumption was completely uncoupled, delta psi generation significantly inhibited, and CsA-dependent delta psi formation prevented. In addition, mitochondria isolated from control cells still underwent swelling, which was partially or completely prevented in mitochondria isolated from apoptotic or necrotic cells, respectively. The moderate swelling was accompanied by a slight rupture of the outer membrane and by a release of cytochrome c. These results point to the involvement of a K(+)ATP channel during the manifestation of PCD induced by H2O2 or NO in plants.

Pea (Pisum sativum) stem mitochondria, energized by NADH, succinate or malate plus glutamate, underwent a spontaneous low-amplitude permeability transition (PT), which could be monitored by dissipation of the electrical potential (deltapsi) or swelling. The occurrence of the latter effects was dependent on O2 availability, because O2 shortage anticipated the manifestation of both deltapsi dissipation and swelling. Spontaneous deltapsi collapse was also monitored in sucrose-resuspended mitochondria and again O2 deprivation caused an anticipation of the phenomenon. However, in this case deltapsi dissipation was not accompanied by a parallel mitochondrial swelling. The latter effect was, indeed, evident only if mitochondria were resuspended in KCl (as osmoticum), or other cations with a molecular mass up to 100 Da (choline+). PT was also induced by protonophores (carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP) or free fatty acids) or valinomycin (only in KCl). The FCCP-induced dissipation of deltapsi and swelling were inhibited by ATP and stimulated (anticipated) by cyclosporin A or O2 shortage. The FCCP-induced PT was accompanied by the release of pyridine nucleotides from the matrix and of cytochrome c from the intermembrane space of KCl-resuspended mitochondria. The spontaneous and FCCP-induced low-amplitude PT of plant mitochondria are interpreted as due to the activity of a recently identified K(ATP)+ channel whose open/closed state is dependent on polarization of the inner membrane and on the oxidoreductive state of some sulfhydryl groups.

A soluble protein with a molecular mass of 55 kDa has been purified from etiolated pea stem mitochondria. The protein exhibits a Mg2+-requiring PPiase activity, with an optimum at pH 9.0, which is not stimulated by monovalent cations, but inhibited by F-, Ca2+, aminomethylenediphosphate and imidodiphosphate. The protein does not cross-react with polyclonal antibodies raised against vacuolar, mitochondrial or soluble PPiases, respectively. Conversely, it cross-reacts with an antibody for the alpha/beta-subunit of the ATP synthase from beef heart mitochondria. The purified protein has been analyzed by MALDI-TOF mass spectrometry and the results, covering the 30% of assigned sequence, indicate that it corresponds to the beta-subunit of the ATP synthase of pea mitochondria. It is suggested that this enzymatic protein may perform a dual function as soluble PPiase or as subunit of the more complex ATP synthase.

The matrix level of pyrophosphate (PPi) in mitochondria isolated from etiolated pea (Pisum sativum L. cv. Alaska) stems was evaluated, on the basis of an enzymatic assay, to be approx. 0.2 mM. Pyrophosphate could enter from the cytoplasm to the mitochondria via adenine nucleotide translocase (ANT), because F- and Ca2+ (two penetrating PPiase inhibitors) and atractylate (ANT inhibitor) inhibited PPiase activity in isolated mitochondria supplied with PPi. This result was also confirmed by measuring oxygen consumption and membrane potential (DeltaPsi) in succinate-energized mitochondria. In a medium free of phosphate (Pi), the addition of PPi before the substrate rendered possible an ADP-stimulated oxygen consumption that was inhibited by F- or Ca2+. In a similar experiment, ADP induced the dissipation of DeltaPsi when it was added after the succinate-generated DeltaPsi had reached a steady state and, again, F- inhibited this dissipation. These results imply that PPi enters the mitochondria where it is hydrolyzed to 2 Pi which become available for the H+-ATPase (EC 3.6.1.34). In addition, PPi may be synthesized by the H+-PPiase (EC 3.6.1.1), acting as a synthase. This evidence arises from the observation that Pi stimulated an oxygen consumption (respiratory control ratio of 1.7) that was inhibited by F- or Ca2+. The physiological role of the mitochondrial H+-PPiase is discussed in the light of the consideration that this enzyme can catalyse a readily reversible reaction.