The toxicity of Cd^^ in vivo during the early phases of radish {Raphanus sativus L.) seed germination and the in vitro Cd^* effect on radish caimodulin (CaM) were studied. Cd^"^ was taken up in the emhryo axes of radish seeds; the increase in fresh weight of emhryo axes after 24 h of incubation was inhihited significantly in the presence of 10 mmol m~^^ Cd^* in the external medium, when the Cd^* content in the embryo axes was c. 1-1 |imol g~' FW. The reabsorption of K*, which characterizes germination, was inhibited by Cd^*, suggesting that Cd^* affected metabolic reactivation. The slight effect of Cd^^ on the transmembrane electric potential of the cortical cells of the emhryo axes excluded a generalized toxicity of Cd^* at the plasma membrane level. After 24 h of incubation, Cd^* induced no increase in total acid-soluble thiols and Cd^*-binding peptides able to reduce Cd^^ toxicity. Ca^* added to the incubation medium partially reversed the Cd^'^-induced inhibition of the increase in fresh weight of embryo axes and concomitantly reduced Cd^* uptake. Equilibrium dialysis experiments indicated that Cd^^ bound to CaM and competed with Ca^^ in this binding. Cd^* inhibited the activation of Ca^^-CaM-dependent calf-brain phosphodiesterase, inhibiting the Ca^^-CaM active complex. Cd^r educed the binding of CaM to the Ca'^^-CaM binding enzymes present in the soluble fraction of the embryo axes of radish seeds. The possibility that Cd^* toxicity in radish seed germination is mediated by the action of Cd^* on Ca *-CaM is discussed in relation to the in vivo and in vitro effects of Cd^K

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Summary• The changes in endopolygalacturonase (endo-PG) levels and endo-PG expression in nonmelting flesh (NMF) and melting flesh (MF) peach fruits ( Prunus persica ) during softening were studied. The endo-PG gene was analysed to identify polymorphisms exploitable for early marker-assisted selection (MAS) of flesh texture.• The role of endo-PG in softening was assessed by western and northern blotting and by biochemical analyses. Polymorphisms in the endo-PG gene were revealed by reverse transcription-polymerase chain reaction (RT-PCR) and sequencing.• An endo-PG protein was detected in both NMF and MF fruits. The levels of this endo-PG protein were higher and increased with softening in MF fruits, but remained lower and were constant in NMF fruits. The different levels of endo-PG appeared to be caused by the differential expression of an endo-PG gene, whose open-reading frame (ORF) showed five single nucleotide polymorphisms (SNPs) in NMF 'Oro A' compared with MF 'Bolero'. One of these SNPs allowed us to determine the allelic configuration at the melting flesh ( M ) locus and also seemed to be exploitable for early MAS in other NMF/MF phenotypes.• The NMF phenotype does not seem to be caused by a large deletion of the endo-PG gene.

Basil (Ocimum basilicum L.) is a culinary, medicinal, and ornamental plant appreciated for its antioxidant properties, mainly attributed to high content of rosmarinic acid. This species also includes purple varieties, characterized by the accumulation of anthocyanins in leaves and flowers. In this work, we compared the main morphological characteristics, the antioxidant capacity and the chemical composition in leaves, flowers, and corollas of green (‘Italiano Classico’) and purple (‘Red Rubin’ and ‘Dark Opal’) basil varieties. The LC-ESI-MS/MS analysis of individual compounds allowed quantifying 17 (poly)phenolic acids and 18 flavonoids, differently accumulated in leaves and flowers of the three varieties. The study revealed that in addition to rosmarinic acid, basil contains several members of the salvianolic acid family, only scarcely descripted in this species, as well as, especially in flowers, simple phenolic acids, such as 4-hydroxybenzoic acid and salvianic acid A. Moreover, the study revealed that purple leaves mainly contain highly acylated anthocyanins, while purple flowers accumulate anthocyanins with low degree of decoration. Overall, this study provides new biochemical information about the presence of not yet characterized bioactive compounds in basil that could contribute to boosting the use of this crop and to gaining new knowledge about the roles of these compounds in plant physiology.

Biostimulants are substances able to improve water and nutrient use efficiency and counteract stress factors by enhancing primary and secondary metabolism. Premise of the work was to exploit raw extracts from leaves (LE) or flowers (FE) of Borago officinalis L., to enhance yield and quality of Lactuca sativa ‘Longifolia,’ and to set up a protocol to assess their effects. To this aim, an integrated study on agronomic, physiological and biochemical aspects, including also a phenomic approach, has been adopted. Extracts were diluted to 1 or 10 mL L–1, sprayed onto lettuce plants at the middle of the growing cycle and 1 day before harvest. Control plants were treated with water. Non-destructive analyses were conducted to assess the effect of extracts on biomass with an innovative imaging technique, and on leaf photosynthetic efficiency (chlorophyll a fluorescence and leaf gas exchanges). At harvest, the levels of ethylene, photosynthetic pigments, nitrate, and primary (sucrose and total sugars) and secondary (total phenols and flavonoids) metabolites, including the activity and levels of phenylalanine ammonia lyase (PAL) were assessed. Moreover, a preliminary study of the effects during postharvest was performed. Borage extracts enhanced the primary metabolism by increasing leaf pigments and photosynthetic activity. Plant fresh weight increased upon treatments with 10 mL L–1 doses, as correctly estimated by multi-view angles images. Chlorophyll a fluorescence data showed that FEs were able to increase the number of active reaction centers per cross section; a similar trend was observed for the performance index. Ethylene was three-fold lower in FEs treatments. Nitrate and sugar levels did not change in response to the different treatments. Total flavonoids and phenols, as well as the total protein levels, the in vitro PAL specific activity, and the levels of PAL-like polypeptides were increased by all borage extracts, with particular regard to FEs. FEs also proved efficient in preventing degradation and inducing an increase in photosynthetic pigments during storage. In conclusion, borage extracts, with particular regard to the flower ones, appear to indeed exert biostimulant effects on lettuce; future work will be required to further investigate on their efficacy in different conditions and/or species.

An inhibitor of Ca2_-calmodulin (Cam)-dependent brain phosphodiesterase was present in the soluble fraction of embryo axes from ungerminated radish (Raphanus sativus L.) seeds. This inhibitor is a Ca2+-dependent, Cam-binding protein; in fact: (a) its effect was strongly reduced by treatment with proteases; (b) the inhibition was counteracted by Cam but not by Ca2 ; (c) on gel filtration in the presence of Ca2+, Cam co-chromatographed with the inhibitor. The inhibitor is heat stable and positively charged at pH 7.5. During early phases of germination, the fresh weight and the levels of DNA and RNA of embryo axes increased, the level of the inhibitor decreased, and the level of Cam increased. Abscisic acid (ABA) inhibited germination, the decrease of inhibitor, and the increase of Cam. Fusicoccin (FC) stimulated the increase in fresh weight but not the increase in the RNA and DNA levels; in this condition, the inhibitor level decreased and the increase in Cam level was higher than in the control. In the presence of both ABA and FC, there was an increase in fresh weight not accompanied by an increase in DNA and RNA levels; Cam increased and, on a fresh weight basis, reached the value of the control. These results indicate that the Ca2+-Cam system was activated in early germination of radish seeds by an increase in Cam and a decrease in the inhibitor levels, that FC, probably through the activation of membrane functions, increased Cam level, and that the ABA inhibition on germination was not mediated by the Ca2+-Cam system. (12), to the maturation of oocytes of Xenopus (6), and to DNA synthesis and mitosis (27). In plants a relationship between Cam level and cell growth is suggested by the fact that higher levels (17-fold) of Cam were found in the root apex (meristem) with respect to the zone of cell elongation (1) and by results on seed germination. Cam level increases during early phases of germination; this increase is reduced when germination is inhibited by ABA, and Cam level appears to be a limiting factor for the Ca2+-Cam-dependent activities (8).An inhibitor of Ca2+-Cam-dependent PDE is present in radish (Raphanus sativus L.) seeds. This inhibitor could be involved in the regulation of Ca2+-Cam-dependent activities and, consequently, of seed germination (8).Seed germination is also characterized by the reactivation of membrane functions (4, 9). FC, a toxin able to stimulate proton extrusion and K+ absorption and to increase the negative electric transmembrane potential (19), stimulates germination, while ABA inhibits germination and the reactivation of membrane functions. FC reverses, in early germination phases, the inhibition of ABA on membrane functions and germination (4).The present paper deals with the evidence that the inhibitor, present in radish seeds, is a Cam-binding protein and reports the changes in Cam and inhibitor levels during early germination phases of radish embryos when germination was stimulated by FC or inhibited by ABA. The role of Cam and of the inhibitor in germination is d...

‘Big Top’ nectarine [Prunus persica (L.) Batsch] has appreciable keeping quality because it resembles, at ripening, the stony hard (SH) peach (P. persica) in firmness and crispness but melts at a slow speed at full ripening. We have characterized the postharvest behavior of ‘Big Top’ fruit, treated or not with ethylene for 5 days after harvest (DAH), and compared it with that of a SH peach (‘Ghiaccio’). Pp-ACS1 expression, ethylene evolution, endo-polygalacturonase (endo-PG) production, and softening were evaluated and compared with those of the physiologically ripe melting flesh (M) cultivar Bolero. Like ‘Bolero’, ‘Big Top’ fruit expressed Pp-ACS1 and evolved ethylene but with a 5-day delay. Pp-endo-PG expression, production of an active endo-PG, and fruit melting showed a parallel behavior; ethylene treatment enhanced all these features. In SH ‘Ghiaccio’ Pp-ACS1 expression, ethylene evolution, endo-PG production, and softening were absent during the first 5 DAH in air. ‘Ghiaccio’ neither expressed Pp-ACS1 nor evolved ethylene even after ethylene treatment but responded by accumulating Pp-endo-PG transcripts and an active endo-PG protein, with flesh melting. A ‘Big Top’ Pp-endo-PG clone showed several single nucleotide (SNP) and insertion-deletion (InDel) polymorphisms in comparison with the M Pp-endo-PG clone of ‘Bolero’ and substantial similarity with the Pp-endo-PG clone of ‘Ghiaccio’. In ‘Big Top’, we identified a peculiar SNP (bp 348) and InDels shared with ‘Ghiaccio’, possibly suitable for discriminating among different genotypes. Overall, the data confirm the pivotal role of ethylene in the regulation of endo-PG production and in the determination of peach flesh texture and support the evidence that ‘Big Top’ could be classified as a melting (slow-melting) phenotype.

Basil (Ocimum basilicum L.) comprises green and purple cultivars, worldwide cultivated and appreciated for high contents of rosmarinic acid and anthocyanins, respectively. Although nitrogen (N) fertilization is needed for high yields, in basil it could have detrimental effects on the accumulation of phenolic compounds. In this study, plants of the cultivars ‘Italiano Classico’ (green) and ‘Red Rubin’ (purple) were grown in hydroponics and subjected to different nutritional treatments, consisting in N starvation, and nitrate (NO3−) or ammonium (NH4+) nutrition. Plant growth and nutritional status, estimated by the contents of NO3−, NH4+, and amino acids in roots and leaves, were evaluated and put in relation with quality traits of basil leaves, such as chlorophyll content, antioxidant capacity, total phenols, the activity of phenylalanine ammonia-lyase, and the concentrations of individual (poly)phenolic acids and flavonoids. This study reveals that N starvation, as well as the availability of the two inorganic N forms, differently affect the phenolic composition in the two cultivars. Compared to plants grown in NO3− availability, in NH4+ availability, green basil showed a higher content of (poly)phenolic acids, while in purple basil, an increase in the contents of anthocyanins was detected. Overall, the study suggests that the management of NH4+ supply could contribute to enhance crop quality in hydroponics, and provides new knowledge about the relationship between N nutrition and phenolic metabolism in basil.