Effects of polyamines on the expression of antioxidant genes and proteins in citrus plants exposed to salt stress

Although there are accumulating reports that polyamines are involved in abiotic/oxidative stress responses, their role is not yet fully understood. Salt stress is one of the most devastating abiotic stresses which seriously interrupt plant growth and productivity. The present study attempts to examine the effects of root treatments with putrescine (Put, I mM), spermidine (Spd, ImM) and spermine (Spm, ImM) on polyamine homeostasis, as well as on several antioxidant-related genes and proteins in the leaves of citrus plants (Citrus aurantium L.) exposed to 150 mM NaCI for 15 d. Analysis of endogenous levels of free polyarnines in NaCl-stressed plant tissues reveals the existence of a polyamine transport system from roots to leaves. Real-time analysis of reactive oxygen species (ROS) by confocal laser scanning microscopy (CLSM) showed an over-accumulation of superoxide anion (02) and hydrogen peroxide (H202) in the stomata of citrus plants exposed to salt stress. Exogenously applied polyamines to salinized nutrient solution induced the activities of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), dehydroascorbate reductase (DHAR) and ascrobic oxidase (AO) whereas it caused the opposite effect on peroxidase (POD), guaiacol peroxidase (GPO D) and ascorbate peroxidase (APX). The effect of polyamines was further examined by determining the plant's antioxidant gene expression profile following a quantitative real-time RT-PCR approach. The overall results indicate that the interaction between different polyamines can be dispersed throughout the citrus plant, and provide additional information suggesting that polyamines may act as a biological mediator allowing citrus plants to activate specific antioxidant responses against salinit

Non-chemical treatments for preventing the postharvest fungal rotting of citrus caused by Penicillium digitatum (green mold) and Penicillium italicum (blue mold)

Background: Citrus is one of the most economically important horticultural crops in the world. Citrus are vulnerable to the postharvest decay caused by Penicillium digitatum and P. italicum, which are both wound pathogens. To date, several non-chemical postharvest treatments have been investigated for the control of both pathogens, trying to provide an alternative solution to the synthetic fungicides (imazalil, thiabendazole, pyrimethanil, and fludioxonil), which are mainly employed and may have harmful effects on human health and environment. Scope and approach: The current study emphasizes the non-chemical postharvest treatments, such as irradiations, biocontrol agents, natural compounds, hot water treatment (HWT), and salts, on the prevention of decay caused by P. digitatum and P. italicum, also known as green and blue molds, respectively. The mode of action of each technique is presented and comprehensively discussed. Key findings and conclusions: In vivo and in vitro experiments in a laboratory scale have shown that the control of green and blue molds can be accomplished by the application of non-chemical treatments. The mechanisms of action of the non-chemical techniques have not been clearly elucidated. Several studies have mentioned that the application of non-chemical treatments results in the synthesis of secondary metabolites with antifungal activities (i.e. polyphenols, phytoalexins) in fruit surface. Moreover, non-chemical treatments may exert direct effects on fungal growth, such as disruption of cell walls, inhibition of metabolic respiration, and disruption of energy production related enzymes.Centro de Investigación y Desarrollo en Criotecnología de Alimento

Principles and Practices in Fruit Tree Production and Postharvest Management

In the near future, the world’s population will face serious challenges due to continuous growth, global climate change, and limited natural resources [...

Η επίδραση του οξειδίου του αζώτου (ΝΟ) και του υδροθείου (H2S) στον εγκλιματισμό σπορόφυτων νεραντζιάς (Citrus aurantium L.) σε συνθήκες ξηρασίας

Nitrosative status has emerged as a key component in plant response to abiotic stress, however, knowledge on its regulation by different environmental conditions remains unclear. In plants, experimental data justify the role of NO as an intercellular signaling molecule under abiotic stress conditions, but data concerning H2S are scarce. The current study focused initially on nitrosative responses in citrus plants exposed to six abiotic stresses using physiological and molecular approaches. Phenotypical observations and molecular analysis showed that abiotic stress treatments were sensed by citrus plants. Furthermore, it was revealed that nitrosative networks are activated by environmental stress factors in citrus leaves as evidenced by increased nitrite content along with the release of NO and superoxide anion (O2-●) in the vascular tissues. The expression of genes potentially involved in NO production, was affected by the abiotic stress treatments demonstrating that NO-derived nitrosative responses could be regulated by various pathways. In addition, GSNOR and NR gene expression and enzymatic activity displayed significant changes in response to adverse environmental conditions, particularly cold stress. The type of abiotic treatment modified differently the ONOO- scavenging ability of citrus plants as well as the protein nitration levels and the protein S-nitrosylation patterns. In a second experiment the effect of SNP and NaHS in the acclimation of citrus plants to drought stress (induced by PEG) was investigated. The roots of citrus plants were pretreated with SNP or NaHS and afterwards exposed to PEG treatments for 21 days. Phenotypical observations and molecular analysis showed that both pre-treatments induced the acclimation of citrus plants to PEG-derived drought stress. Under drought stress the portions of nitrite and NOx were increased, but decreased under SNP or NaHS pre-treatment. Under drought stress, total RSNOs levels were modified and a significant modulation in GSNOR activity was observed following both pre-treatments. The activity of LCD was changed under drought condition, whereas H2S level was changed by both pre-treatments under drought. Several changes in carbonylation and nitration levels were recorded in plant pre-treated with SNP or NaHS. Also, NO or H2S, caused an increase in S-nitrosylation levels under water stress. Following mass spectrometry analysis 33 S-nitrosylated proteins were identified which mainly are involved in photosynthesis, photorespiration and abiotic defense. Also, via proteomic analysis several different protein patterns were observed among treatments, providing clues for specific protein alternations driven by drought, SNP or H2S. These results suggest that the nitrosative response of citrus plants is differentially regulated depending on the stress type and underscore the possible role of GSNOR as a potent molecular marker of cold stress in citrus. Overall these data support the existence of an interaction among NO and H2S during the acclimation of citrus plants to drought stress.Η γνώση των επιπέδων των ενεργών μορφών αζώτου (RNS) αποτελεί κλειδί στη μελέτη των αβιοτικών καταπονήσεων. Πτυχές του μηχανισμούς δράσης των RNS κατά την διάρκεια των τελευταίων ετών δεν έχουν διευκρινιστεί πλήρως. Στα φυτά, πειραματικά δεδομένα τεκμηριώνουν τον ρόλο του ΝΟ ως ενδοκυτταρικού χημικού αγγελιοφόρου, ενώ ο ρόλος του H2S είναι υπό διερεύνηση. Η παρούσα εργασία μελετά αρχικά τις αποκρίσεις φυτών νεραντζιάς (Citrus aurantium L.) ως προς τα επίπεδα μεταβολής των RNS έπειτα από την έκθεση των φυτών σε έξι αβιοτικές καταπονήσεις. Με ανατομικές, μοριακές, βιοχημικές μετρήσεις, διαπιστώθηκε ότι οι αβιοτικές καταπονήσεις οδήγησαν στην αύξηση της συγκέντρωσης των NO2-, με παράλληλη απελευθέρωση ΝΟ και Ο2-● στον αγωγό ιστό. Επίσης, οι διάφορες αβιοτικές καταπονήσεις μετέβαλαν την έκφραση γονιδίων που σχετίζονται με την παραγωγή του ΝΟ. Σε συνθήκες ψύχους, η έκφραση του γονιδίου αλλά και η δράση του ενζύμου της GSNOR αλλά και της NR μεταβλήθηκε. Επίσης οι αβιοτικές καταπονήσεις επηρέασαν την ικανότητα εκκαθάρισης του ΟΝΟΟ-. Τα επίπεδα της νίτρωσης και της S-νιτροσυλίωσης των πρωτεϊνών μεταβλήθηκαν μεταξύ των αβιοτικών καταπονήσεων. Στη συνέχεια o πειραματισμός επικεντρώθηκε στην προσπάθεια επαγωγής του εγκλιματισμού των φυτών με την προ-μεταχείρισή τους με ΝΟ ή H2S. Ανατομικές, μοριακές, βιοχημικές μετρήσεις κατέδειξαν την θετική απόκριση των φυτών στην προ-μεταχείριση με ΝΟ ή H2S. Η ποσότητα των NO2- και ΝΟx αυξήθηκε σε συνθήκες υδατικής καταπόνησης, ενώ μειώθηκε στις προ-μεταχειρίσεις με ΝΟ ή H2S. Σε συνθήκες ξηρασίας παρατηρήθηκε μεταβολή στην συγκέντρωσης των ολικών RSNO και σημαντική τροποποίηση της δράσης της GSNOR όταν τα φυτά επωάστηκαν σε διαλύματα ΝΟ ή H2S. Σε συνθήκες υδατικής καταπόνησης, επηρεάστηκε η δράση της LCD και διαπιστώθηκε τροποποίηση της συγκέντρωσης H2S, όταν τα φυτά προ-μεταχειρίστηκαν με ΝΟ ή H2S. Η γονιδιακή έκφραση της NR, σε συνθήκες υδατικής καταπόνησης παρέμεινε στα επίπεδα του μάρτυρα όταν τα φυτά προ-μεταχειρίστηκαν με ΝΟ ή H2S. Επίσης, σε συνθήκες υδατικής καταπόνησης, μειώθηκε η καρβονυλίωση και η νίτρωση των πρωτεϊνών όταν στα φυτά προ-μεταχειρίστηκαν με ΝΟ ή H2S. Επίσης, η προ-μεταχείριση με ΝΟ ή με H2S αύξησε την S-νιτροσυλίωση. Η ανάλυση των S-νιτροσυλιωμένων πρωτεϊνών έδειξε ότι 33 πρωτεΐνες S-νιτροσυλιώθηκαν, οι οποίες στη συνέχεια διαπιστώθηκε ότι εμπλέκονται κυρίως στη φωτοσύνθεση. Επίσης, με ανάλυση του πρωτεϊνικού χάρτη στα φύλλα εντοπίστηκαν οι πρωτεϊνικές μεταβολές ως προς την υδατική καταπόνηση και τις προ-μεταχειρίσεις με ΝΟ ή με H2S. Με βάση τα αποτελέσματα προτείνεται ότι το είδος της καταπόνησης καθορίζει τον τρόπο δράσης των RNS. Η GSNOR δύναται να αποτελέσει βιοχημικό δείκτη της καταπόνησης ψύχους στα εσπεριδοειδή. Σε συνθήκες υδατικής καταπόνησης, τα πειραματικά δεδομένα τεκμηριώνουν την ύπαρξη αλληλεπίδρασης μεταξύ των ΝΟ και H2S. Σε συνθήκες υδατικής καταπόνησης, το ΝΟ και το H2S επιδρά σε εξειδικευμένους πρωτεϊνικούς στόχους, ενισχύοντας τον ρόλο των συγκεκριμένων βιομορίων στον εγκλιματισμό σε περιβαλλοντικές καταπονήσεις

Involvement of AsA/DHA and GSH/GSSG Ratios in Gene and Protein Expression and in the Activation of Defence Mechanisms Under Abiotic Stress Conditions

In a persistently changing environment there are many adverse abiotic stress conditions such as cold, heat, drought, salinity, heavy metal toxicity and oxygen deprivation, which remarkably influence plant growth and crop production. Plant cells produce oxygen radicals and their derivatives, so-called reactive oxygen species (ROS) during various processes associated with abiotic stress. Moreover, the generation of ROS is the main means for higher plants to transmit cellular signalling information concerning the changing environmental conditions. Therefore, plants have evolved inducible redox state-based sensing mechanisms that are activated or amplified in response to adverse environmental conditions. Ascorbate and glutathione, the key cellular redox buffers, are used for both detoxification of ROS and transmission of redox signals. In recent years, it has become clear that abiotic stress conditions induce changes in the reduction/oxidation (redox) state of signalling molecules, which in turn modulate gene and protein expression to increase plant acclimation to abiotic stress. This important redox state-related branch of science has given several clues in understanding the adaptive plant responses to different stressful regimes. In this chapter, an overview of the literature is briefly presented in terms of the main function of ascorbate and glutathione in plant cells. Further more, we describe how important forms of abiotic stress regulate the expression of genes and proteins involved in the ascorbate and glutathione redox sensing system

Hydrogen Sulfide: A Potent Tool in Postharvest Fruit Biology and Possible Mechanism of Action

Hydrogen sulfide (H2S), an endogenous gaseous molecule, is considered as a signaling agent, in parallel with other low molecular weight reactive substances, mainly hydrogen peroxide (H2O2) and nitric oxide (NO), in various plant systems. New studies are now revealing that the postharvest application of H2S, through H2S donors such as sodium hydrosulfide (NaSH) or sodium sulfide (Na2S), can inhibit fruit ripening and senescence programs in numerous fruits. We discuss here current knowledge on the impact of H2S in postharvest physiology of several climacteric and non-climacteric fruits such as banana, apple, pear, kiwifruit, strawberry, mulberry fruit, and grape. Although there is still a considerable lack of studies establishing the mechanisms by which H2S signaling is linked to fruit metabolism, we highlight several candidate mechanisms, including a putative cross-talk between H2S and ethylene, reactive oxygen and nitrogen species, oxidative/nitrosative stress signaling, sulfate metabolism, and post-translational modification of protein cysteine residues (S-sulfhydration) as being functional in this H2S postharvest action. Understanding H2S metabolism and signaling during postharvest storage and the interplay with other key player molecules would therefore provide new, improved strategies for better fruit postharvest storage. To achieve this understanding, postharvest fruit physiology research will need to focus increasingly on the spatial interaction between H2S and ethylene perception as well as on the interplay between S-sulfhydration/desulfhydration and S-nitrosylation/denitrosylation under several postharvest conditions

Preharvest Foliar Application of Si–Ca-Based Biostimulant Affects Postharvest Quality and Shelf-Life of Clementine Mandarin (Citrus clementina Hort. Ex Tan)

Citriculture and the postharvest industry are in the quest for biostimulants that favour fruit quality and extend shelf-life. Recently, Si has emerged as a biostimulant and its impact on fruit quality and postharvest shelf-life needs to be elucidated. The experiment is conducted for two consecutive years (2019 and 2020) in a commercial citrus orchard. In the present study, a Si–Ca-based product (Gravital® Force SC, AGROLOGY SA, Sindos, Greece) is foliar sprayed upon clementine mandarin (Citrus clementina Hort. Ex Tan cv. SRA 63) trees from August to November, while unsprayed trees are kept as controls. At commercial maturity, both sprayed and unsprayed fruits are harvested and stored for thirty (30) days at 5 °C with 90–95% relative humidity. Afterwards, they are kept at shelf temperature (20 °C) for six (6) days (shelf-life). At different intervals [at harvest, after cold storage (30 d at 5 °C), at the third day of shelf-life (30 d at 5 °C plus 3 d at 20 °C) and sixth day of shelf-life (30 d at 5 °C plus 6 d at 20 °C)], fruits are sampled and analysed for their qualitative characteristics. According to the results, the preharvest foliar application of the Si–Ca-based product delayed fruit maturation, increased peel firmness, total soluble content, total acidity, ascorbic acid, total phenols and antioxidant capacity, and reduced fruit decay during shelf storage. Results suggest that the preharvest foliar spray of Si–Ca products is able to maintain the postharvest quality of mid-ripening mandarin fruit

Preharvest Foliar Application of Si–Ca-Based Biostimulant Affects Postharvest Quality and Shelf-Life of Clementine Mandarin (<i>Citrus clementina</i> Hort. Ex Tan)

Citriculture and the postharvest industry are in the quest for biostimulants that favour fruit quality and extend shelf-life. Recently, Si has emerged as a biostimulant and its impact on fruit quality and postharvest shelf-life needs to be elucidated. The experiment is conducted for two consecutive years (2019 and 2020) in a commercial citrus orchard. In the present study, a Si–Ca-based product (Gravital® Force SC, AGROLOGY SA, Sindos, Greece) is foliar sprayed upon clementine mandarin (Citrus clementina Hort. Ex Tan cv. SRA 63) trees from August to November, while unsprayed trees are kept as controls. At commercial maturity, both sprayed and unsprayed fruits are harvested and stored for thirty (30) days at 5 °C with 90–95% relative humidity. Afterwards, they are kept at shelf temperature (20 °C) for six (6) days (shelf-life). At different intervals [at harvest, after cold storage (30 d at 5 °C), at the third day of shelf-life (30 d at 5 °C plus 3 d at 20 °C) and sixth day of shelf-life (30 d at 5 °C plus 6 d at 20 °C)], fruits are sampled and analysed for their qualitative characteristics. According to the results, the preharvest foliar application of the Si–Ca-based product delayed fruit maturation, increased peel firmness, total soluble content, total acidity, ascorbic acid, total phenols and antioxidant capacity, and reduced fruit decay during shelf storage. Results suggest that the preharvest foliar spray of Si–Ca products is able to maintain the postharvest quality of mid-ripening mandarin fruit

Polyamines modify the nitrosative status of salt stressed plants

Although there is accumulating evidence that polyamines are involved in the response of plants to both salinity and nitrosative stress, their role is not yet fully understood. The present study attempts to examine the effects of hydroponic root treatments with putrescine (Put, 1 mM), spermidine (Spd, 1mM) and spermine (Spm, 1mM) on nitrosative homeostasis in leaves of citrus plants (Citrus aurantium L.) exposed to 100 mM NaCI for 15 d. Nitric oxide (NO) steady-state levels and DAF-2DA-derived fluorescence were stimulated by NaCI treatment and especially by the application of polyamines in the salt-treated plants. Transcriptional analysis showed that the expression of several genes encoding proteins associated with NO biosynthesis, including AOX, XO, GSNO, NOS, NiR and NR, is regulated by polyamine application. In addition, S-nitrosoglutathione reductase (GSNOR) activity was suppressed in plants treated with Put and Spd. The profile of tyrosine-nitrated proteins was diminished by the application of polyamines. The characterization of nitroproteome by mass fingerprinting revealed several protein targets which are involved mainly in photosynthesis, disease/defense, energy and protein destination/storage. These data strongly support a function of polyamines in modulating the nitrosative status upon salt stress