Publications by authors named "Roberto De Michele"

Posidonia oceanica meadows are the most productive coastal ecosystem in the Mediterranean. Posidonia oceanica seeds are enclosed in buoyant fleshy fruits that allow dispersal. Many fruits eventually strand on beaches, imposing a remarkable energy cost for the plant.

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Understanding mechanisms of nutrient allocation in organisms requires precise knowledge of the spatiotemporal dynamics of small molecules in vivo. Genetically encoded sensors are powerful tools for studying nutrient distribution and dynamics, as they enable minimally invasive monitoring of nutrient steady-state levels in situ. Numerous types of genetically encoded sensors for nutrients have been designed and applied in mammalian cells and fungi.

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Robust protocols for the regeneration of somatic embryos in vitro are essential for the efficient use of the most modern biotechnologies. Unfortunately, in perennial trees such as , plants regenerated from juvenile tissues usually exhibit strong, undesirable juvenile characters such as thorny habit and delayed flowering and fruit production. In this work, we tested whether the cell types (nucellar and stigma/style) used to regenerate plants through somatic embryogenesis affected the transition from the juvenile to mature phase.

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Fumonisin B induces rapid programmed cell death in Arabidopsis cells, oxidative and nitrosative bursts, and differentially modulates cell death responsive genes. Glutathione is the main antioxidant involved in the stress response. Fumonisin B (FB) is a fungal toxin produced by Fusarium spp.

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Grapevine ( ssp. ) is a perennial crop especially important for wine and fruit production. The species is highly polymorphic with thousands of different varieties selected by farmers and clonally propagated.

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Heavy metal pollution is an increasing global concern. Among heavy metals, mercury (Hg) is especially dangerous because of its massive release into the environment and high toxicity, especially for aquatic organisms. The molecular response mechanisms of algae to Hg exposure are mostly unknown.

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Caper (Capparis spinosa L.) is a xerophytic shrub cultivated for its flower buds and fruits, used as food and for their medicinal properties. Breeding programs and even proper taxonomic classification of the genus Capparis has been hampered so far by the lack of reliable genetic information and molecular markers.

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Hydrogen sulfide (HS) has been recently found as an important signaling molecule especially in root system architecture of plants. The regulation of root formation through HS has been reported in previous works; while the profiling of metabolites in response to HS is not clearly discussed. To this end, different concentrations of sodium hydrosulfide (an HS donor) were applied to the culture of Linum album hairy roots.

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Sensitivity, dynamic and detection range as well as exclusion of expression and instrumental artifacts are critical for the quantitation of data obtained with fluorescent protein (FP)-based biosensors in vivo. Current biosensors designs are, in general, unable to simultaneously meet all these criteria. Here, we describe a generalizable platform to create dual-FP biosensors with large dynamic ranges by employing a single FP-cassette, named GO-(Green-Orange) Matryoshka.

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The plant plasma membrane is the interface between the cell and its environment undertaking a range of important functions related to transport, signaling, cell wall biosynthesis, and secretion. Multiple proteomic studies have attempted to capture the diversity of proteins in the plasma membrane using biochemical fractionation techniques. In this study, two-phase partitioning was combined with free-flow electrophoresis to produce a population of highly purified plasma membrane vesicles that were subsequently characterized by tandem mass spectroscopy.

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We recently described a series of genetically encoded, single-fluorophore-based sensors, termed AmTrac and MepTrac, which monitor membrane transporter activity in vivo (De Michele et al., 2013). However, being intensiometric, AmTrac and Meptrac are limited in their use for quantitative studies.

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Leaf senescence is the last stage of development of an organ and is aimed to its ordered disassembly and nutrient reallocation. Whereas chlorophyll gradually degrades during senescence in leaves, mitochondria need to maintain active to sustain the energy demands of senescing cells. Here we analysed the motility and morphology of mitochondria in different stages of senescence in leaves of grapevine (Vitis vinifera), by stably expressing a GFP (green fluorescent protein) reporter targeted to these organelles.

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Biosensors offer an innovative tool for measuring the dynamics of a wide range of metabolites in living organisms. Biosensors are genetically encoded, and thus can be specifically targeted to specific compartments of organelles by fusion to proteins or targeting sequences. Mitochondria are central to eukaryotic cell metabolism and present a complex structure with multiple compartments.

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Ammonium is a preferred source of nitrogen for plants but is toxic at high levels. Plant ammonium transporters (AMTs) play an essential role in NH4(+) uptake, but the mechanism by which AMTs are regulated remains unclear. To study how AMTs are regulated in the presence of ammonium, we used variable-angle total internal reflection fluorescence microscopy and fluorescence cross-correlation spectroscopy for single-particle fluorescence imaging of EGFP-tagged AMT1;3 on the plasma membrane of Arabidopsis root cells at various ammonium levels.

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Ammonium serves as key nitrogen source and metabolic intermediate, yet excess causes toxicity. Ammonium uptake is mediated by ammonium transporters, whose regulation is poorly understood. While transport can easily be characterized in heterologous systems, measuring transporter activity in vivo remains challenging.

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Ammonium and urea are important nitrogen sources for autotrophic organisms. Plant genomes encode several families of specific transporters for these molecules, plus other uptake mechanisms such as aquaporins and ABC transporters. Selaginella and Physcomitrella are representatives of lycophytes and bryophytes, respectively, and the recent completion of their genome sequences provided us with an opportunity for comparative genome studies, with special emphasis on the adaptive processes that accompanied the conquest of dry land and the evolution of a vascular system.

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S-Nitrosoglutathione (GSNO) is a bioactive, stable, and mobile reservoir of nitric oxide (NO), and an important player in defence responses to herbivory and pathogen attack in plants. It has been demonstrated previously that GSNO reductase (GSNOR) is the main enzyme responsible for the in vivo control of intracellular levels of GSNO. In this study, the role of S-nitrosothiols, in particular of GSNO, in systemic defence responses in Arabidopsis thaliana was investigated further.

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Studies performed in different organisms have highlighted the importance of protein kinase CK2 in cell growth and cell viability. However, the plant signaling pathways in which CK2 is involved are largely unknown. We have reported that a dominant-negative mutant of CK2 in Arabidopsis thaliana shows phenotypic traits that are typically linked to alterations in auxin-dependent processes.

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Protein kinase CK2 is a pleiotropic Ser/Thr kinase, evolutionary conserved in eukaryotes. Studies performed in different organisms, from yeast to humans, have highlighted the importance of CK2 in cell growth and cell-cycle control. However, the signalling pathways in which CK2 is involved have not been fully identified.

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Senescence is the last stage of development of a leaf, and is marked by a progressive degradation of cellular components and their recycling to other organs of a plant. We recently performed a transcriptomic analysis of leaf senescence in the legume model . Changes in gene expression reflected the profound physiological rearrangement and provided intriguing clues for the involvement of some of the members identified, which await future investigation.

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Exposure to cadmium (Cd(2+)) can result in cell death, but the molecular mechanisms of Cd(2+) cytotoxicity in plants are not fully understood. Here, we show that Arabidopsis (Arabidopsis thaliana) cell suspension cultures underwent a process of programmed cell death when exposed to 100 and 150 microm CdCl(2) and that this process resembled an accelerated senescence, as suggested by the expression of the marker senescence-associated gene12 (SAG12). CdCl(2) treatment was accompanied by a rapid increase in nitric oxide (NO) and phytochelatin synthesis, which continued to be high as long as cells remained viable.

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Here, for the first time, a comprehensive transcriptomics study is presented of leaf senescence in the legume model Medicago truncatula, providing a broad overview of differentially expressed transcripts involved in this process. The cDNA-amplification fragment length polymorphism (AFLP) technique was used to identify > 500 genes, which were cloned and sorted into functional categories according to their gene ontology annotation. Comparison between the datasets of Arabidopsis and M.

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The relationship between nitric oxide (NO) and salicylic acid (SA) was investigated in Arabidopsis thaliana. Here it is shown that SA is able to induce NO synthesis in a dose-dependent manner in Arabidopsis. NO production was detected by confocal microscopic analysis and spectrofluorometric assay in plant roots and cultured cells.

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Nitric oxide (NO) is a gaseous free radical that reacts with O(2) in air and aqueous solution. NO donors have been widely used to circumvent the difficulties inherent in working with a reactive gas, but NO donors do not deliver NO at a constant rate for prolonged periods of time. Furthermore, some of the most commonly used NO donors produce additional, bioactive decomposition products.

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