Sheng Shu

Effects of exogenous spermine on chlorophyll fluorescence, antioxidant system and ultrastructure of chloroplasts in Cucumis sativus L. under salt stress.

The effects of exogenous spermine (Spm) on plant growth, chlorophyll fluorescence, ultrastructure and anti-oxidative metabolism of chloroplasts were investigated in Cucumis sativus L. under NaCl stress. Salt stress significantly reduced plant growth, chlorophylls content and F(v)/F(m). These changes could be alleviated by foliar spraying with Spm. Salt stress caused an increase in malondialdehyde (MDA) content and superoxide anion [Formula: see text] generation rate in chloroplasts. Application of Spm significantly increased activities of superoxidase dismutase (SOD, EC 1.15.1.1), peroxidase (POD, EC 1.11.1.7), and ascorbate peroxidase (APX, EC 1.11.1.11) which decreased the levels of [Formula: see text] and MDA in the salt-stressed chloroplasts. Salt stress decreased the activities of dehydroascorbate reductase (DHAR, EC 1.8.5.1) and glutathione reductase (GR, EC 1.6.4.2) in the chloroplasts and reduced the contents of dehydroascorbate (DAsA) and glutathione (GSH), but increased monodehydroascorbate reductase (MDAR, EC 1.6.5.4) activity. On the other hand, Spm significantly increased the activities of antioxidant enzymes and levels of antioxidants in the salt-stressed chloroplasts. Further analysis of the ultrastructure of chloroplasts indicated that salinity induced destruction of the chloroplast envelope and increased the number of plastoglobuli with aberrations in thylakoid membranes. However, Spm application to salt-stressed plant leaves counteracted the adverse effects of salinity on the structure of the photosynthetic apparatus. These results suggest that Spm alleviates salt-induced oxidative stress through regulating antioxidant systems in chloroplasts of cucumber seedlings, which is associated with an improvement of the photochemical efficiency of PSII.

Effects of salt stress on the structure and function of the photosynthetic apparatus in Cucumis sativus and its protection by exogenous putrescine

With the objective to clarify the physiological significance of polyamines (PAs) in the photosynthetic apparatus, the present study investigated the effects of salt stress with and without foliar application of putrescine (Put) on the structure and function of the photosynthetic apparatus in cucumber. Salt stress at 75 mM NaCl for 7 days resulted in a severe reduction of photosynthesis. The fast chlorophyll afluorescence transient analysis showed that salt stress inhibited the maximum quantum yield of PSII photochemistry (F(v)/F(m)), mainly due to damage at the receptor side of PSII. In addition, salt stress decreased the density of active reaction centers and the structure performance. The microscopic analysis revealed that salt stress-induced destruction of the chloroplast envelope and increased the number of plastoglobuli along with aberrations in thylakoid membranes. Besides, salt stress caused a decrease in the content of endogenous PAs, conjugated and bound forms of spermidine and spermine in particular, in thylakoid membranes. However, applications of 8 mM Put alleviated the salt stress-mediated decrease in net photosynthetic rates (Pn) and actual efficiency of PSII(Φ(PSII)). Put increased PAs in thylakoid membranes and overcame the damaging effects of salt stress on the structure and function of the photosynthetic apparatus in salt-stressed plant leaves. Put application to control plants neither increased PAs in thylakoid membranes nor affected photosynthesis. These results indicate that PAs in chloroplasts play crucial roles in protecting the thylakoid membranes against the deleterious influences of salt stress. In addition, the present results point to the probability that the salt-induced dysfunction of photosynthesis is largely attributable to the loss of PAs in the photosynthetic apparatus.

Effects of proline on photosynthesis, root reactive oxygen species (ROS) metabolism in two melon cultivars ( Cucumis melo L.) under NaCl stress

Effects of 0.2 mM proline applied to saline nutrient solution on biomass, chlorophyll content, photosynthetic parameters, reactive oxygen species and antioxidant enzymes activities of two melon cultivars (cv. Yuhuang and cv. Xuemei) were examined. Results indicate that exogenous proline increased the fresh and dry weights of both melon cultivars under NaCl stress, raised their chlorophyll content, net photosynthetic rate (Pn), actual efficiency of photosystem II (ΦPSII), enhanced the activity of SOD, POD, CAT, APX, DHAR and GR in their roots, lowered the superoxide anion radical level and reduced the hydrogen peroxide (H 2 O 2 ) content and malondialdehyde (MDA) content. Exogenous proline also alleviate salinity-induced damage of membrane in both melon cultivars. In conclusion, proline treatment enhanced the salinity tolerance of both melon plants and alleviated their salinity-induced damage. However, all the above effects of proline were markedly more significant in cv. Xuemei than in cv. Yuhuang, suggesting that proline had different effects on different cultivars of melon plants. Key words : Proline, photosynthesis, reactive oxygen species, antioxidant enzymes, salt tolerance, NaCl stress.

The role of putrescine in the regulation of proteins and fatty acids of thylakoid membranes under salt stress.

Polyamines can alleviate the inhibitory effects of salinity on plant growth by regulating photosynthetic efficiency. However, little information is available to explain the specific mechanisms underlying the contribution of polyamines to salt tolerance of the photosynthetic apparatus. Here, we investigated the role of putrescine (Put) on the photosynthetic apparatus of cucumber seedlings under salt stress. We found that NaCl stress resulted in severe ion toxicity and oxidative stress in cucumber chloroplasts. In addition, salinity caused a significant increase in the saturated fatty acid contents of thylakoid membranes. Put altered unsaturated fatty acid content, thereby alleviating the disintegration of thylakoid grana lamellae and reducing the number of plastoglobuli in thylakoid membranes. BN-PAGE revealed Put up-regulated the expression of ATP synthase, CP47, D1, Qb, and psbA proteins and down-regulated CP24, D2, and LHCII type III in NaCl-stressed thylakoid membranes. qRT-PCR analysis of gene expression was used to compare transcript and protein accumulation among 10 candidate proteins. For five of these proteins, induced transcript accumulation was consistent with the pattern of induced protein accumulation. Our results suggest that Put regulates protein expression at transcriptional and translational levels by increasing endogenous polyamines levels in thylakoid membranes, which may stabilise photosynthetic apparatus under salt stress.

Effects of exogenous spermidine on photosynthetic capacity and expression of Calvin cycle genes in salt-stressed cucumber seedlings

We investigated the effects of exogenous spermidine (Spd) on growth, photosynthesis and expression of the Calvin cycle-related genes in cucumber seedlings (Cucumis sativus L.) exposed to NaCl stress. Salt stress reduced net photosynthetic rates (PN), actual photochemical efficiency of PSII (ΦPSII) and inhibited plant growth. Application of exogenous Spd to salinized nutrient solution alleviated salinity-induced the inhibition of plant growth, together with an increase in PN and ΦPSII. Salinity markedly reduced the maximum carboxylase activity of ribulose-1,5-bisphosphate carboxylase/oxygenase (Vcmax), the maximal velocity of RuBP regeneration (Jmax), triose-phosphate utilization capacity (TPU) and carboxylation efficiency (CE). Spd alleviated the negative effects on CO2 assimilation induced by salt stress. Moreover, Spd significantly increased the activities and contents of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) and fructose-1,6-biphosphate aldolase (ALD; aldolase) in the salt-stressed cucumber leaves. On the other hand, salinity up-regulated the transcriptional levels of ribulose-1,5-bisphosphate (RCA), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoribrokinase (PRK) and down-regulated the transcriptional levels of ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit (RbcL), ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit (RbcS), ALD, triose-3-phosphate isomerase (TPI), fructose-1,6-bisphosphate phosphatase (FBPase) and 3-phosphoglyceric acid kinase (PGK). However, Spd application to salt-stressed plant roots counteracted salinity-induced mRNA expression changes in most of the above-mentioned genes. These results suggest that Spd could improve photosynthetic capacity through regulating gene expression and activity of key enzymes for CO2 fixation, thus confers tolerance to salinity on cucumber plants.

A Review: Polyamines and Photosynthesis

Polyamines (PAs) are low molecular weight ubiquitous nitrogenous compounds found in all living organisms (Kaur-Sawhney et al., 2003). In higher plants, the most common polyamines are spermidine (Spd), spermine (Spm) and their diamine obligate precursor putrescine (Put). They are formed by aliphatic hydrocarbons substituted with two or more amino groups (Figure.1). Because of the polycationic nature at physiological pH, PAs are present in the free form or as conjugates bound to phenolic acids and other low molecular weight compounds or to proteins and nucleic acids (Childs et al., 2003). Like hormones, PAs displaying high biological activity are involved in a wide array of fundamental processes in plants, such as replication and gene expression, growth and development, senescence, membrane stabilization, enzyme activity modulation and adaptation to abiotic and biotic stresses (Galston et al., 1997; Bais and Ravishankar, 2002; Zapata et al., 2008). Although, according to these reports, PAs seem to be important growth regulators, their precise physiological function and mechanism of action still remain unclear. It has been shown that chloroplasts and photosynthetic subcomplexes including thylakoids, LHCII complex and PSII membranes are enriched with three major polyamines, while PSII core and the reaction center of PSII are exclusively rich in Spm (Kotzabasis et al., 1993; Navakoudis et al., 2003). The potential role of polyamines in maintaining the photochemical efficiency of plants has become a research focus. These studies mainly focused on the effect PAs exert a positive role in the photosynthesis of plants in response to various environmental stresses. In green alga, it was shown that the bound Put content of the thylakoid membrane was increased in environments with high CO2 concentrations, which caused an increase in reaction center density and led to an increased photosynthetic rate (Logothetis et al., 2004). An increase in conjugated Put content can stabilize the thylakoid membrane, thus enhancing resistance of tobacco plants to ozone pollution (Navakoudis et al., 2003) and UV-B radiation (Lutz et al., 2005). Low temperature stress reduced the content of Put as well as the Put/Spm ratio in thylakoids and the light-harvesting complexes LHCII in Phaseolus vulgaris L., leading to a decrease in photosynthetic electron transport rate and inactivation of the PSII reaction center (Sfakianaki et al., 2006). Put is also involved in the induction of a photosynthetic apparatus owning high concentration of reaction center with a small functional antenna that leads to enhance photochemical quenching of the absorbed light energy (Kotzabasis et al., 1999). PAs biosynthesis is controlled by light and the Spm/Put ratio is correlated to the structure and function of the photosynthetic apparatus

Effects of 24-epibrassinolide on ascorbate–glutathione cycle and polyamine levels in cucumber roots under Ca(NO3)2 stress

The effects of 24-epibrassinolide (EBL) applied by spraying the leaves of Cucumis sativus L (cv. Jinyou No.4) on the roots of ascorbate–glutathione (AsA–GSH) cycle and levels and forms of polyamines were investigated under Ca(NO3)2 stress. Ca(NO3)2 stress caused significant decreases in ascorbate acid (AsA) and glutathione (GSH) levels and the activities of antioxidant enzymes involved in the AsA–GSH cycle, including ascorbate peroxidase (APX, EC 1.11.1.11), glutathione reductase (GR, EC 1.6.4.2), monodehydroascorbate reductase (MDAR, EC 1.6.5.4) and dehydroascorbate reductase (DHAR, EC 1.6.5.4). In contrast, exogenous EBL in NB combination increased AsA and GSH levels through enhancing activities of APX, GR, MDAR and DHAR. This alleviated the declines of AsA/DHA and GSH/GSSG ratios, which could maintain cellular homeostasis. In addition, EBL application reduced the accumulation of putrescine (Put) induced by excess Ca(NO3)2, and promoted the conversion of Put into spermidine (Spd) and/or spermine (Spm) able to bind to cell membrane structures. These results showed that EBL could alleviate the damage from oxidative stress by up-regulating the capacity of the AsA–GSH cycle and altering polyamine levels and forms.

Effects of exogenous spermidine on antioxidant system of tomato seedlings exposed to high temperature stress

The effects of foliar spraying with spermidine (Spd) on antioxidant system in tomato (Lycopersicon esculentum Mill.) seedlings were investigated under high temperature stress. The high temperature stress significantly inhibited plant growth and reduced chlorophyll (Chl) content. Application of exogenous 1 mM Spd alleviated the inhibition of growth induced by the high temperature stress. Malondialdehyde (MDA), hydrogen peroxide (H2O2) content and superoxide anion (O2) generation rate were significantly increased by the high temperature stress, but Spd significantly reduced the accumulation of reactive oxygen species (ROS) and MDA content under the stress. The high temperature stress significantly decreased glutathione (GSH) content and activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX), glutathione reductase (GR), monodehydroascorbate reductase (MDHAR) and dehydroascorbate reductase (DHAR), but increased contents of dehydroascorbic acid (DHA), ascorbic acid (AsA), and oxidized glutathione (GSSG) in tomato leaves. However, Spd significantly increased the activities of antioxidant enzymes, levels of antioxidants and endogenous polyamines in tomato leaves under the high temperature stress. In addition, to varying degrees, Spd regulated expression of MnSOD, POD, APX2, APX6, GR, MDHAR, DHAR1, and DHAR2 genes in tomato leaves exposed to the high temperature stress. These results suggest that Spd could change endogenous polyamine levels and alleviate the damage by oxidative stress enhancing the non-enzymatic and enzymatic antioxidant system and the related gene expression.

The role of 24-epibrassinolide in the regulation of photosynthetic characteristics and nitrogen metabolism of tomato seedlings under a combined low temperature and weak light stress.

In the present study, we aimed to investigate the effects of exogenous 24-epibrassinolide (EBR) on growth, photosynthetic characteristics, chlorophyll fluorescence imaging, and nitrogen metabolism of tomato leaves under low temperature and weak light conditions. The results showed that foliar application of EBR significantly alleviated the inhibition of plant growth, and increased the fresh and dry weights of tomato plants under a combined low temperature and weak light stress. Moreover, EBR also increased the net photosynthetic rate (Pn), light saturation point (LSP), maximal quantum yield of PSII photochemistry (Fv/Fm), actual photochemical efficiency of PSII (ФPSII), and photochemical quenching coefficient (qP), but decreased the intercellular CO2 concentration (Ci), light compensation point (LCP) and apparent quantum efficiency (AQE) under low temperature and weak light conditions. In addition, application of EBR to tomato leaves significantly enhanced the activities of nitrate reductase (NR), glutamate dehydrogenase (GDH), glutamine synthetase (GS), and glutamate synthase (GOGAT), but decreased the ammonium content and nitrite reductase (NiR) activity. We observed that EBR remarkably increased the contents of aspartic acid, threonine, serine, glycine, and phenylalanine, while decreasing the accumulation of cysteine, methionine, arginine, and proline under a combined low temperature and light stress. These results suggest that EBR could alleviate the combined stress-induced harmful effects on photosynthesis and nitrogen metabolism, thus leading to improved plant growth.

Proteomic and Physiological Analyses Reveal Putrescine Responses in Roots of Cucumber Stressed by NaCl

Soil salinity is a major environmental constraint that threatens agricultural productivity. Different strategies have been developed to improve crop salt tolerance, among which the effects of polyamines have been well-reported. To gain a better understanding of the cucumber (Cucumis sativus L.) responses to NaCl and unravel the underlying mechanism of exogenous putrescine (Put) alleviating salt-induced damage, comparative proteomic analysis was conducted on cucumber roots treated with NaCl, and/or Put for 7 days. The results showed that exogenous Put restored the root growth inhibited by NaCl. Sixty-two differentially expressed proteins implicated in various biological processes were successfully identified by MALDI-TOF/TOF MS. The four largest categories included proteins involved in defense response (24.2%), protein metabolism (24.2%), carbohydrate metabolism (19.4%), and amino acid metabolism (14.5%). Exogenous Put up-regulated most identified proteins involved in carbohydrate metabolism, implying an enhancement in energy generation. Proteins involved in defense response and protein metabolism were differently regulated by Put, which indicated the roles of Put in stress resistance and proteome rearrangement. Put also increased the abundance of proteins involved in amino acid metabolism. Meanwhile, physiological analysis showed that Put could further up-regulated the levels of free amino acids in salt stressed-roots. In addition, Put also improved endogenous polyamines contents by regulating the transcription levels of key enzymes in polyamine metabolism. Taken together, these results suggest that Put may alleviate NaCl-induced growth inhibition through degradation of misfolded/damaged proteins, activation of stress defense, and the promotion of carbohydrate metabolism to generate more energy.