Search for compounds able to modulate fof1 atp synthase in switching from life enzyme to cell death executor

Mitochondria are the main site of energy power in eukaryotic cells. The enzyme FOF1 synthase is responsible for ATP production driven by the transmembrane proton gradient. The maintenance of a very low permeability of the inner mitochondrial membrane is crucial for this mechanism, since sudden opening of the permeability transition pore (PTP) leads to matrix swelling and outer membrane rupture, with release of proapoptotic factors. Recently, it has been suggested that dimers of ATP synthase in mammals could represent the main component of the mitochondrial PTP, a feature modulated by calcium and involving the matrix protein Cyclophilin D (CyPD). This study would help to develop new tools for the identification of plant secondary metabolites, in particular flavonoids, able to modulate PTP and therefore acting on the programmed cell death mediated by mitochondria. Therefore, this project would represent the first screening for plant molecules able to interfere with programmed cell death, as a preliminary study for the development of drugs active in PTPrelated pathologies

Caspase-3-like activity and proteasome degradation in grapevine suspension cell cultures undergoing silver-induced programmed cell death

Toxic metal contamination is one of the major environmental concerns of the recent decade, due to the large application of metals in industrial, healthcare and commercial products, even in the form of nanostructures and nanomaterials. Nevertheless, the effects of silver (Ag+) on plants have not yet thoroughly elucidated. Therefore, suspension cell cultures of grapevine were used as a model for investigating silver toxicity. To do this, oxidative stress and programmed cell death (PCD), evaluated as reactive oxygen species production, caspase-3-like activity and ubiquitin-proteasome system, were investigated. As a result, the highest concentration (10 \u3bcM) of Ag+ caused a rapid (within 24 h) induction of PCD (approx. 80%), accompanied by generation of reactive oxygen species and activation of caspase-3-like activity. In the presence of specific inhibitor of this enzyme, a partial recovery of cell viability and a strong inhibition of caspase- 3-like activity was observed. In addition, silver-induced PCD was accompanied either by increase of poly-ubiquitin conjugated proteins and degradation of subunit PBA1 of the proteasome 20S core, similarly to what found for metal-induced neurotoxicity in animals. The present study shows that silver could induce PCD in grapevine suspension cell cultures, mediated by caspase-3-like activity and oxidative stress. These effects were associated to accumulation of poly-ubiquitin conjugated proteins, suggesting the impairment of ubiquitin-proteasome complex, confirmed by the decrease of the PBA1 subunit. These findings indicate that animal and plant cells could share a common pathway in response to toxic metal, which involves PCD and disassembling of proteasome complex

Structural and functional properties of plant mitochondrial F-ATP synthase

The mitochondrial F-ATP synthase is responsible for coupling the transmembrane proton gradient, generated through the inner membrane by the electron transport chain, to the synthesis of ATP. This enzyme shares a basic architecture with the prokaryotic and chloroplast ones, since it is composed of a catalytic head (F1), located in the mitochondrial matrix, a membrane-bound part (FO), together with a central and a peripheral stalk. In this review we compare the structural and functional properties of F-ATP synthase in plant mitochondria with those of yeast and mammals. We also present the physiological impact of the alteration of F-ATP synthase in plants, with a special regard to its involvement in cytoplasmic male sterility. Furthermore, we show the involvement of this enzyme in plant stress responses. Finally, we discuss the role of F-ATP synthase in shaping the curvature of the mitochondrial inner membrane and in permeability transition pore formation

Produção de resinas poliésteres com diferentes monômeros para aplicação em compósitos poliméricos

Este trabalho visa obter compósitos de matriz de resina poliéster insaturada com fibra de vidro com propriedades térmicas e mecânicas semelhantes ao compósito de resina epóxi já consolidada no mercado. Resinas poliéster possuem valor comercial de até um terço do custo das resinas epóxi, porém tem sua aplicabilidade limitada em função da alta contração durante o processo de cura, que pode chegar a 10% do volume de peça. Neste trabalho, foi realizada a avaliação das propriedades da resina poliéster após a reação radicalar com diferentes monômeros como estireno e metacrilato de metila (MMA), com o intuito de avaliar uma possível substituição de compósitos epóxi empregados em áreas do mercado que exigem alto desempenho mecânico como a da aviação.This work aims to obtain composites of unsaturated polyester matrices containing glass fiber with thermal and mechanical properties similar to a an already consolidated epoxy resin composite commercially available. Polyester resins usually cost up to a third of the price of epoxy resins, however they have limited applicability due to the high contraction during the curing process, which can reach 10% of the volume. The properties of the polyester resin were evaluated after the addition of different monomers such as styrene and methyl methacrylate in order to evaluate a possible replacement of epoxy composites used in applications that require high mechanical performance such as aviation

Relationships between population traits, nonstructural carbohydrates, and elevation in alpine stands of Vaccinium myrtillus

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\u2013plant 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

Biochemical and immunochemical similarities among mammalian bilitranslocase and a plant flavonoid translocator

Flavonoids are a large class of plant secondary metabolites, belonging to polyphenol family, which possess pharmacological and nutritional properties. Their synthesis takes place only in plants, while mammals can acquire them only with diet. It has been demonstrated that flavonoid uptake occurs in rat also by the activity of bilitranslocase, a carrier that is involved in anion transport in liver cell, vascular endothelium and gastric mucosa. A sequence of bilitranslocase interacting with flavonoid moieties is already known and characterized. Antibody raised against such protein epitope were shown to exhibit cross-reactivity against plant membrane proteins in tissues involved in flavonoid transport and accumulation, such as teguments of carnation petals and skin of grape berries. Further immunolocalization studies allowed to demonstrate the presence of cross-reacting protein not only at the level of tegumental tissues, but also associated to sieve elements and seed teguments in grape berries

Flavonoids and darkness lower PCD in senescing Vitis vinifera suspension cell cultures

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

CHARACTERIZATION OF LIPASE ACTIVITY IN GREEN COFFEE BEANS DURING STORAGE AND GERMINATION

Lipase, green coffee, germination, storage, lipid Coffee seeds possess an intermediate storage pattern, showing a non-quiescent behaviour characterized by various metabolic reactions occurring during storage. In particular, lipase is the main enzyme involved in the mobilization of triacylglycerols, providing energy and a source of carbon skeleton during early stages of germination. During storage, the triacylglycerols might be involved in the generation of undesirable compounds (known as \u201coff-flavours\u201d), lowering both the viability of coffee seeds and the cup quality. In this work, the soluble protein fraction from coffee seeds and plantlets was extracted by acetone and then was utilised to assay lipase activity. Green coffee beans, harvested in Colombia, were stored at room temperature for 2-3 months, in order to verify the influence of prolonged storage on lipase activity. For germination experiments, the seeds were imbibed for 7 days at 30 \ub0C and transferred in perlite at 28 \ub0C and 90% R.H. for further 3 weeks. Lipase activity was detected by a colorimetric method based on specific degradation of a chromogenic substrate, at pH 8.2. Green coffee seeds exhibited an appreciable lipase activity that was slightly increased during storage. Such an activity was inhibited by tetrahydrolipstatin (THL) in a concentration-dependent manner, while it was slightly stimulated by both EGTA and EDTA. During the germination, after 10, 14, 17 and 21 days, lipase activity showed an initial increase that was followed by a gradual decrease. The effect of the presence or absence of the parchment (seed coat), during the first stages of germination, has also been investigated