Roberto De Michele

Nitric Oxide Is Involved in Cadmium-Induced Programmed Cell Death in Arabidopsis Suspension Cultures

Exposure to cadmium (Cd2+) can result in cell death, but the molecular mechanisms of Cd2+ 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 μm CdCl2 and that this process resembled an accelerated senescence, as suggested by the expression of the marker senescence-associated gene12 (SAG12). CdCl2 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. Hydrogen peroxide production was a later event and preceded the rise of cell death by about 24 h. Inhibition of NO synthesis by NG-monomethyl-arginine monoacetate resulted in partial prevention of hydrogen peroxide increase, SAG12 expression, and mortality, indicating that NO is actually required for Cd2+-induced cell death. NO also modulated the extent of phytochelatin content, and possibly their function, by S-nitrosylation. These results shed light on the signaling events controlling Cd2+ cytotoxicity in plants.

Transcriptome analysis of Medicago truncatula leaf senescence: similarities and differences in metabolic and transcriptional regulations as compared with Arabidopsis, nodule senescence and nitric oxide signalling

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. truncatula leaf senescence reveals common physiological events but differences in the nitrogen metabolism and in transcriptional regulation. In addition, it was observed that a minority of the genes regulated during leaf senescence were equally involved in other processes leading to programmed cell death, such as nodule senescence and nitric oxide signalling. This study provides a wide transcriptional profile for the comprehension of key events of leaf senescence in M. truncatula and highlights a possible regulative role for MADS box transcription factors in the terminal phases of the process.

Legume leaf senescence: A transcriptional analysis

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 Medicago truncatula. 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. The resulting data also revealed similarities and differences with Arabidopsis leaf senescence.

Mitochondria Change Dynamics and Morphology during Grapevine Leaf Senescence

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. Results show that mitochondria were less dynamic and markedly changed morphology during senescence, passing from the elongated, branched structures found in mature leaves to enlarged and sparse organelles in senescent leaves. Progression of senescence in leaves was not synchronous, since changes in mitochondria from stomata were delayed. Mitochondrial morphology was also analysed in grapevine cell cultures. Mitochondria from cells at the end of their growth curve resembled those from senescing leaves, suggesting that cell cultures might represent a useful model system for senescence. Additionally, senescence-associated mitochondrial changes were observed in plants treated with high concentrations of cytokinins. Overall, morphology and dynamics of mitochondria might represent a reliable senescence marker for plant cells.

Intra-varietal genetic diversity of the grapevine ( Vitis vinifera L.) cultivar ‘Nero d’Avola’ as revealed by microsatellite markers

The Sicilian grape cultivar ‘Nero d’Avola’ is among the oldest and most cultivated in the island, taking part in the production of several red wines exported worldwide, including DOC wines (Etna Rosso and Cerasuolo di Vittoria). Due to the ancient origin and repeated clonally propagation of the cultivar, phenotypic variability has been observed. Clone identification in this important cultivar has so far relied on phenotypic and chemical traits analyses, often affected by environmental conditions. Genetic markers, such as microsatellites, are particularly useful for cultivar identification, parentage testing, pedigree reconstruction and population structure studies. In the present paper, microsatellites were used to analyze the intra-varietal genetic diversity among 118 plants of ‘Nero d’Avola’, collected in 30 vineyards displaced in different areas of Sicily. Out of 22 microsatellites, 11 showed polymorphism among samples and 15 different phylogenetic groups were identified. Results show that ‘Nero d’Avola’ actually comprises different genetic profiles, although most of clones share a common origin.

Urgent need for preservation of grapevine (Vitis vinifera L. subsp. vinifera) germplasm from small circum-Sicilian islands as revealed by SSR markers and traditional use investigations

Since the last decades grapevine germplasm is undergoing a process of rapid genetic erosion. This process is of particular concern in minor circum-Sicilian islands, because of the sharp reduction of the cultivated surfaces and the shift of their economy from agriculture to tourism. Aiming at valorising and preserving the surviving varieties we collected 185 accessions during several surveys since 2007. Six nuclear microsatellite markers were used for germplasm characterization, yielding 75 different genetic profiles. We found out that most genetic profiles (39) were not listed in national and international grapevine databases, confirming that the Sicilian minor islands represent underexplored hotspots of genetic diversity for grapevine. We also identified several synonymies, often due to geographic isolation, having 20 varieties at least two names. Conversely, 18 homonyms collectively indicated 34 genetically different accessions. Interviews with farmers provided information on current and past usage, and the origin and type of cultivation practices as well. The study also shows the urgent need for preservation of local grapevine germplasm, due to the disappearance of the elder caretakers of these traditional varieties. For rare germplasm preservation most part of the collected grapevine cultivars were introduced in an ex situ collection field.