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Basi biochimico-molecolari della tolleranza a condizioni di stress osmo-salino in varietà italiane di riso
No full textDifferential accumulation of γ–aminobutyric acid in elicited cells of two rice cultivars showing contrasting sensitivity to the blast pathogen.
No full textIntracellular free amino acid pools were quantified in suspension cultured cells of a blast-sensitive and a blast-resistant rice genotype at increasing time after the treatment with Magnaporthe oryzae cell wall hydrolysates. Besides some expected variations in free phenylalanine, a remarkable early increase of γ–aminobutyric acid (GABA) levels was evident in both cultivars. Glutamate decarboxylase activity and protein levels were unaffected. GABA homeostasis was recovered in the sensitive cultivar 48 h after the treatment. On the contrary, a further GABA accumulation and a general increase of most amino acids were found at this later stage in the resistant genotype, which showed a higher decrease of cell viability as a consequence of elicitor addition. Data support a recently-hypothesized role of GABA metabolism in the plant response to fungal pathogens
Intraspecific variability of floral nectar volume and composition in rapeseed (Brassica napus L. var. oleifera)
No full textNumerous angiosperms rely on pollinators to ensure efficient flower fertilization, offering a reward consisting of nourishing nectars produced by specialized floral cells, known as nectaries. Nectar components are believed to derive from phloem sap that is enzymatically processed and transformed within nectaries. An increasing body of evidence suggests that nectar composition, mainly amino acids, may influence pollinator attraction and fidelity. This notwithstanding, little is known about the range of natural variability in nectar content for crop species. Sugar and amino acid composition of nectar harvested from field-grown plants at the 63 to 65 phenological stage was determined for a set of 44 winter genotypes of rapeseed, a bee-pollinated crop. Significant differences were found for solute concentrations, and an even higher variability was evident for nectar volumes, resulting in striking differences when results were expressed on a single flower basis. The comparison of nectar and phloem sap from a subset of 8 varieties pointed out qualitative and quantitative diversities with respect to both sugars and amino acids. Notably, amino acid concentration in phloem sap was up to one hundred times higher than in nectar. Phloem sap showed a much more uniform composition, suggesting that nectar variability depends mainly on nectary metabolism. A better understanding of the basis of nectar production would allow an improvement of seed set efficiency, as well as hive management and honey production
Amino acid content and nectar choice by forager honeybees (Apis mellifera L.)
No full textDual choice feeding tests were performed to determine a preference of forager honeybees for specific amino acids. Artificial nectar containing proline was preferred over that containing only sugars. Nectar containing alanine was preferred on the first day, but preference was no longer significant thereafter. On the contrary, a negative response was found for serine. When the bees were given the choice between two nectars enriched with different compounds, proline was preferred above both alanine and serine, and alanine above serine
Divergent properties and phylogeny of 5-enol-pyruvyl-shikimate-3-phosphate synthases from cyanobacteria: evidence supporting horizontal gene transfer in the Nostocales
No full textAs it represents the target of the successful herbicide glyphosate, great attention has been paid to the shikimate pathway enzyme 5-enol-pyruvyl-shikimate-3-phosphate (EPSP) synthase. However, inconsistent results have been reported concerning the sensitivity of the enzyme from cyanobacteria, and consequent inhibitory effects on cyanobacterial growth. The properties of EPSP synthase were investigated in a set of 42 strains representative of the large morphological diversity of these prokaryotes. Publicly available protein sequences were analyzed, and related to enzymatic features. In most cases, the native protein showed an unusual homodimeric composition and a general sensitivity to micromolar doses of glyphosate. By contrast, eight out of 15 Nostocales strains were found to possess a monomeric EPSP synthase, whose activity was inhibited only at concentrations exceeding 1mM. Sequence analysis showed that these two forms are only distantly related, the latter clustering separately in a clade composed of diverse bacterial phyla. The results are consistent with the occurrence of a horizontal gene transfer event involving an evolutionarily distant organism. Moreover, data suggest that the existence of class I (glyphosate-sensitive) and class II (glyphosate-tolerant) EPSP synthases representing two distinct phylogenetic clades is an oversimplification because of the limited number of analyzed samples
Sublethal detergent concentrations increase metabolization of recalcitrant polyphosphonates by the cyanobacterium Spirulina platensis
No full textAs a consequence of increasing industrial applications, thousand tons of polyphosphonates are introduced every year into the environment. The inherent stability of the C–P bond results in a prolonged half-life. Moreover, low uptake rates limit further their microbial metabolization. To assess whether low detergent concentrations were able to increase polyphosphonate utilization by the cyanobacterium Spirulina platensis, tolerance limits to the exposure to various detergents were determined by measuring the growth rate in the presence of graded levels below the critical micellar concentration. Then, the amount of hexamethylenediamine-N,N,N′,N′-tetrakis(methylphosphonic acid) that is metabolized in the absence or in the presence of sublethal detergent concentrations was quantified by 31P NMR analysis on either P-starved or P-fed cyanobacterial cultures. The strain tolerated the presence of detergents in the order: nonionic > anionic > cationic. When added to the culture medium at the highest concentrations showing no detrimental effects upon cell viability, detergents either improved or decreased polyphosphonate
utilization, the anionic sodium dodecyl sulfate being the most
beneficial. Metabolization was not lower in P-fed cells—a
result that strengthens the possibility of using, in the future, this strain for bioremediation purposes
Synthesis and evaluation of effective inhibitors of plant δ1-pyrroline-5-carboxylate reductase
No full textAnalogues of previously studied phenyl-substituted aminomethylene-bisphosphonic acids were synthesized and
evaluated as inhibitors of Arabidopsis thaliana δ1-pyrroline-5-carboxylate reductase. With the aim of improving their effectiveness, two main modifications were introduced into the inhibitory scaffold: the aminomethylenebisphosphonic moiety was replaced with a hydroxymethylenebisphosphonic group, and the length of the molecule was increased by replacing the methylene linker with an ethylidene chain. In addition, chlorine atoms in the phenyl ring were replaced with various other substituents. Most of the studied derivatives showed activity in the micromolar to millimolar range, with two of them being more effective than the lead compound, with concentrations inhibiting 50% of enzyme activity as low as 50 μM. Experimental evidence supporting the ability of these inhibitors to interfere with proline synthesis in vivo is also shown
Phytotoxicity of aminobisphosphonates targeting both δ1-pyrroline-5-carboxylate reductase and glutamine synthetase
No full textBACKGROUND: Dual-target inhibitors may contribute to the management of herbicide-resistant weeds and avoid or delay the selection of resistant biotypes. Some aminobisphosphonates inhibit the activity of both glutamine synthetase and δ1-pyrroline-5-carboxylate (P5C) reductase in vitro, but the relevance of the latter in vivo has yet to be proven. This study aimed at demonstrating that these compounds can also block proline synthesis in planta. RESULTS: Two aminophosphonates, namely 3,5-dichlorophenylamino-methylenebisphosphonic acid and 3,5-dibromophenylaminomethylenebis phosphonic acid (Br2PAMBPA), showed inverse effectiveness against the two partially purified target enzymes from rapeseed. The compounds showed equipotency in inhibiting the growth of rapeseed seedlings and cultured cells. The analysis of amino acid content in treated cells showed a strong reduction in glutamate and glutamate-related amino acid pools, but a milder effect on free proline. In the case of Br2PAMBPA, toxic P5C levels accumulated in treated seedlings, proving that the inhibition of P5C reductase takes place in situ. CONCLUSIONS: Phenyl-substituted aminobisphosphonates may be regarded as true dual-target inhibitors. Their use to develop new active principles for crop protection could consequently represent a tool to address the problem of target-site resistance among weeds
A differential tolerance to mild salt stress conditions among six Italian rice genotypes does not rely on Na+ exclusion from shoots
No full textRice is very sensitive to salt stress at the seedling level, with consequent poor crop establishment. A natural variability in susceptibility to moderate saline environments was found in a group of six Italian temperate japonica rice cultivars, and the physiological determinants for salt tolerance were investigated. Cation (Na+, K+ and Mg++) levels were determined in shoots from individual rice plantlets grown in the absence or in the presence of inhibitory, yet sublethal salt levels, and at increasing time after salt treatments. Significant variations were found among genotypes, but these were unrelated to the relative tolerance, which seems to result from neither mechanism(s) for reduced Na+ translocation to the aerial part, nor its increased retrieval from the xylem mediating Na+ exclusion from leaves. Accordingly, thiobarbituric acid reactive substance levels raised in leaf tissues of salt-treated seedlings, and osmo-induced proline accumulation was found in all genotypes. Data suggest that the difference in salt tolerance most likely depends on mechanisms for osmotic adjustment and/or antioxidative defence
Functional characterization and expression analysis of rice δ1-pyrroline-5-carboxylate dehydrogenase provide new insight into the regulation of proline and arginine catabolism
No full textWhile intracellular proline accumulation in response to various stress conditions has been investigated in great detail, the biochemistry and physiological relevance of proline degradation in plants is much less understood. Moreover, the second and last step in proline catabolism, the oxidation of δ1-pyrroline-5-carboxylic acid (P5C) to glutamate, is shared with arginine catabolism. Little information is available to date concerning the regulatory mechanisms coordinating these two pathways. Expression of the gene coding for P5C dehydrogenase was analyzed in rice by real-time PCR either following the exogenous supply of amino acids of the glutamate family, or under hyperosmotic stress conditions. The rice enzyme was heterologously expressed in E. coli, and the affinity-purified protein was thoroughly characterized with respect to structural and functional properties. A tetrameric oligomerization state was observed in size exclusion chromatography, which suggests a structure of the plant enzyme different from that shown for the bacterial P5C dehydrogenases structurally characterized to date. Kinetic analysis accounted for a preferential use of NAD+ as the electron acceptor. Cations were found to modulate enzyme activity, whereas anion effects were negligible. Several metal ions were inhibitory in the micromolar range. Interestingly, arginine also inhibited the enzyme at higher concentrations, with a mechanism of uncompetitive type with respect to P5C. This implies that millimolar levels of arginine would increase the affinity of P5C dehydrogenase towards its specific substrate. Results are discussed in view of the involvement of the enzyme in either proline or arginine catabolism
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