Remodeling of chloroplast proteome under salinity affects salt tolerance of <Emphasis Type="Italic">Festuca arundinacea</Emphasis>

Acclimation of photosynthetic apparatus to variable environmental conditions is an important component of tolerance to dehydration stresses, including salinity. The present study deals with the research on alterations in chloroplast proteome of the forage grasses. Based on chlorophyll fluorescence parameters, two genotypes of a model grass species—Festuca arundinacea with distinct levels of salinity tolerance: low salt tolerant (LST) and high salt tolerant (HST), were selected. Next, two-dimensional electrophoresis and mass spectrometry were applied under both control and salt stress conditions to identify proteins accumulated differentially between these two genotypes. The physiological analysis revealed that under NaCl treatment the studied plants differed in photosystem II activity, water content, and ion accumulation. The differentially accumulated proteins included ATPase B, ATP synthase, ribulose-1,5-bisphosphate carboxylase large and small subunits, cytochrome b6-f complex iron-sulfur subunit, oxygen-evolving enhancer proteins (OEE), OEE1 and OEE2, plastidic fructose-bisphosphate aldolase (pFBA), and lipocalin. A higher level of lipocalin, potentially involved in prevention of lipid peroxidation under stress, was also observed in the HST genotype. Our physiological and proteomic results performed for the first time on the species of forage grasses clearly showed that chloroplast metabolism adjustment could be a crucial factor in developing salinity tolerance.     

<Emphasis Type="Bold">Role of microRNAs and their target genes in salinity response in plant</Emphasis>s

We examined the effects of 2,4-epibrassinolide (EBR) application on photosynthesis, antioxidant enzyme activity, and Rubisco activase (RCA) gene expression in wheat (Triticum aestivum L.) seedlings under a combination of drought and heat stress. The net photosynthetic rates (P_n) of wheat seedlings decreased significantly, the photosynthetic capability was inhibited, and the activities of superoxide (SOD), peroxidase (POD), catalase (CAT), and RCA as well as the initial and total activity of Rubisco declined under the combined stress. These decreases and inhibitory effects were significantly ameliorated by exogenous EBR application. Three subunits (45–46, 41–42, and 38–39 kDa) of RCA were observed in wheat seedlings. The abundances of the 38–39 kDa and 41–42 kDa subunits were significantly lower in plants subjected to stressful conditions than in unstressed plants. Interestingly, a marked increase in 45–46 kDa RCA was observed under heat or heat combined with drought stress. The abundance of 38–39 kDa RCA in seedlings exposed to heat, drought, or their combination was significantly enhanced by EBR pretreatment, which paralleled the changes in initial Rubisco activity and P_n, but was not consistent with observed mRNA abundance. These results indicated that the larger subunit of RCA (45–46 kDa), which is more thermostable and increased in response to moderate heat stress, and the smaller isoform (38–39 kDa) of RCA may play important roles in maintaining the photosynthetic capability by EBR under stress conditions.     

Effects of genetic manipulation of the activity of photorespiration on the redox state of photosystem I and its robustness against excess light stress under CO<Subscript>2</Subscript>-limited conditions in rice

Under CO₂-limited conditions such as during stomatal closure, photorespiration is suggested to act as a sink for excess light energy and protect photosystem I (PSI) by oxidizing its reaction center chlorophyll P700. In this study, this issue was directly examined with rice (Oryza sativa L.) plants via genetic manipulation of the amount of Rubisco, which can be a limiting factor for photorespiration. At low [CO₂] of 5 Pa that mimicked stomatal closure condition, the activity of photorespiration in transgenic plants with decreased Rubisco content (RBCS-antisense plants) markedly decreased, whereas the activity in transgenic plants with overproduction of Rubisco (RBCS-sense plants) was similar to that in wild-type plants. Oxidation of P700 was enhanced at [CO₂] of 5 Pa in wild-type and RBCS-sense plants. PSI was not damaged by excess light stress induced by repetitive saturated pulse-light (rSP) in the presence of strong steady-state light. On the other hand, P700 was strongly reduced in RBCS-antisense plants at [CO₂] of 5 Pa. PSI was also damaged by rSP illumination. These results indicate that oxidation of P700 and the robustness of PSI against excess light stress are hampered by the decreased activity of photorespiration as a result of genetic manipulation of Rubisco content. It is also suggested that overproduction of Rubisco does not enhance photorespiration as well as CO₂ assimilation probably due to partial deactivation of Rubisco.     

Photoinhibition of photosystem I in <Emphasis Type="Italic">Nephrolepis falciformis</Emphasis> depends on reactive oxygen species generated in the chloroplast stroma

We studied how high light causes photoinhibition of photosystem I (PSI) in the shade-demanding fern Nephrolepis falciformis, in an attempt to understand the mechanism of PSI photoinhibition under natural field conditions. Intact leaves were treated with constant high light and fluctuating light. Detached leaves were treated with constant high light in the presence and absence of methyl viologen (MV). Chlorophyll fluorescence and P700 signal were determined to estimate photoinhibition. PSI was highly oxidized under high light before treatments. N. falciformis showed significantly stronger photoinhibition of PSI and PSII under constant high light than fluctuating light. These results suggest that high levels of P700 oxidation ratio cannot prevent PSI photoinhibition under high light in N. falciformis. Furthermore, photoinhibition of PSI in N. falciformis was largely accelerated in the presence of MV that promotes the production of superoxide anion radicals in the chloroplast stroma by accepting electrons from PSI. From these results, we propose that photoinhibition of PSI in N. falciformis is mainly caused by superoxide radicals generated in the chloroplast stroma, which is different from the mechanism of PSI photoinhibition in Arabidopsis thaliana and spinach. Here, we provide some new insights into the PSI photoinhibition under natural field conditions.     

Protection of photosystem I during sudden light stress depends on ferredoxin:NADP(H) reductase abundance and interactions

Plant tolerance to high light and oxidative stress is increased by overexpression of the photosynthetic enzyme Ferredoxin:NADP(H) reductase (FNR), but the specific mechanism of FNR-mediated protection remains enigmatic. It has also been reported that the localization of this enzyme within the chloroplast is related to its role in stress tolerance. Here, we dissected the impact of FNR content and location on photoinactivation of photosystem I (PSI) and photosystem II (PSII) during high light stress of Arabidopsis (Arabidopsis thaliana). The reaction center of PSII is efficiently turned over during light stress, while damage to PSI takes much longer to repair. Our results indicate a PSI sepcific effect, where efficient oxidation of the PSI primary donor (P700) upon transition from darkness to light, depends on FNR recruitment to the thylakoid membrane tether proteins: thylakoid rhodanase-like protein (TROL) and translocon at the inner envelope of chloroplasts 62 (Tic62). When these interactions were disrupted, PSI photoinactivation occurred. In contrast, there was a moderate delay in the onset of PSII damage. Based on measurements of ΔpH formation and cyclic electron flow, we propose that FNR location influences the speed at which photosynthetic control is induced, resulting in specific impact on PSI damage. Membrane tethering of FNR therefore plays a role in alleviating high light stress, by regulating electron distribution during short-term responses to light.

Altered location of a key enzyme involved in the post-photosystem I electron transport chain ameliorates damage to photosystem I during increasing light intensity.     

Exogenously-supplied trehalose protects thylakoid membranes of winter wheat from heat-induced damage

The effects of trehalose pretreatment on thylakoid membranes of winter wheat were investigated under heat stress. Under normal growth conditions, the winter wheat synthesized 502 μg g⁻¹(f.m.) trehalose, which increased to 1250 μg g⁻¹(f.m.) under heat stress and to 1658 μg g⁻¹(f.m.) in trehalose-pretreated seedlings. Under heat stress, proteins in the thylakoid membranes and the photosynthetic capacity were protected by trehalose pretreatment. Moreover, the electrolyte leakage, contents of malondialdehyde, superoxide anion and hydrogen peroxide, and lipoxygenase activity in trehalose-pretreated seedlings were lower than in the non-pretreated plants.     

Photosynthetic response of tetraploid and hexaploid wheat to water stress

Photosynthetic characteristics of ear and flag leaves of wheat species, tetraploid Triticum dicoccoides Kom and hexaploid Bima1, were studied in plants grown under well-watered (WW) and water-stressed (WS) conditions. Compared to ears, flag leaves exhibited higher photosynthetic rate (P _N) at the filling stage, but more severe decrease under WS. P _N in the tetraploid wheat ear remained higher than that in the hexaploid wheat during the grain-filling stage. Water stress decreased PN in both the organs; this decline was caused by a reduction in Rubisco activity, not by drought-induced stomatal limitation. Tetraploid wheat ears exhibited higher relative water content and water-use efficiency than that of hexaploid wheat, under WS. The change in phosphoenolpyruvate carboxylase activity and carbon isotope composition indicated the absence of C₄ metabolism in the ears of both species under both conditions. The improved performance of the tetraploid wheat ears under WS was associated with better water relations.     

Induction of water deficit tolerance in wheat due to exogenous application of plant growth regulators: membrane stability,water relations and photosynthesis

Our experiment was carried out in order to explore effects of plant growth regulators (PGR; thidiazuron, paclobutrazol, and ascorbic acid) on physiological traits of wheat genotypes under water surplus and deficit conditions. Study revealed that relative water content, membrane stability index, chlorophyll content, photosynthetic rate (P_N), and maximal quantum yield of PSII improved with PGRs application across the genotypes both under irrigation and water stress. The response of HD 2733 genotype was more positive toward PGRs treatment as compared to other genotypes under water stress. Higher P_N and chlorophyll contents were observed in HD 2987 followed by C 306 genotype under water-stress conditions. Moreover, Rubisco small subunit (SSU) expression was lower in wheat genotypes under water stress as compared to irrigated conditions. Application of PGRs led to upregulation of SSU under water stress, while no significant change was found in Rubisco level and activity under irrigated condition in dependence on PGRs treatments. Yield-related traits showed also significant reduction under water-stress conditions, while application of PGRs enhanced the yield and its components. Results indicated that the PGRs exhibited a positive interaction and synergetic effect on water stressed wheat plants in terms of photosynthetic machinery and yield.     

Involvement of small heat shock proteins,trehalose, and lipids in the thermal stress management in <Emphasis Type="Italic">Schizosaccharomyces pombe</Emphasis>

Changes in the levels of three structurally and functionally different important thermoprotectant molecules, namely small heat shock proteins (sHsps), trehalose, and lipids, have been investigated upon heat shock in Schizosaccharomyces pombe. Both α-crystallin-type sHsps (Hsp15.8 and Hsp16) were induced after prolonged high-temperature treatment but with different kinetic profiles. The shsp null mutants display a weak, but significant, heat sensitivity indicating their importance in the thermal stress management. The heat induction of sHsps is different in wild type and in highly heat-sensitive trehalose-deficient (tps1Δ) cells; however, trehalose level did not show significant alteration in shsp mutants. The altered timing of trehalose accumulation and induction of sHsps suggest that the disaccharide might provide protection at the early stage of the heat stress while elevated amount of sHsps are required at the later phase. The cellular lipid compositions of two different temperature-adapted wild-type S. pombe cells are also altered according to the rule of homeoviscous adaptation, indicating their crucial role in adapting to the environmental temperature changes. Both Hsp15.8 and Hsp16 are able to bind to different lipids isolated from S. pombe, whose interaction might provide a powerful protection against heat-induced damages of the membranes. Our data suggest that all the three investigated thermoprotectant macromolecules play a pivotal role during the thermal stress management in the fission yeast.     

Photosynthetic responses of the low intertidal macroalga Sargassum fusiforme (Sargassaceae) to saline stress

Sargassum fusiforme, a species of brown seaweed with economic importance, inhabits lower intertidal zones where algae are often exposed to various stresses. In this study, changes in the photosynthetic performance of S. fusiforme under saline stress were investigated. The PSII performance in S. fusiforme significantly improved, when the thalli were exposed to 0% salinity, and remained high with prolonging treatment time. In contrast, the PSII activity declined considerably under salinities of 4.5 and 6%. The PSI activity did not change remarkably under saline stress, thus demonstrating higher tolerance to saline stress than PSII. In addition, the PSI activity could be also restored after saline treatments, when PSII was inhibited by 3-(3,4-dichlorophenyl)-1,1-dimethylurea. It might be as a result of changes in the NAD(P)H content in the thalli under saline stress. Our results suggested that PSI was much more tolerant to different saline stress than PSII in S. fusiforme. We demonstrated that S. fusiforme was much more tolerant to hyposaline than to hypersaline stress.     

Over-Expression of <Emphasis Type="Italic">PmHSP17.9</Emphasis> in Transgenic <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> Confers Thermotolerance

Small heat shock proteins (sHSPs) have been shown to be involved in stress tolerance. However, their functions in Prunus mume under heat treatment are poorly characterized. To improve our understanding of sHSPs, we cloned a sHSP gene, PmHSP17.9, from P. mume. Sequence alignment and phylogenetic analysis indicated that PmHSP17.9 was a member of plant cytosolic class III sHSPs. Besides heat stress, PmHSP17.9 was also upregulated by salt, dehydration, oxidative stresses and ABA treatment. Leaves of transgenic Arabidopsis thaliana that ectopically express PmHSP17.9 accumulated less O₂ ^? and H₂O₂ compared with wild type (WT) after 42 °C treatment for 6 h. Over-expression of PmHSP17.9 in transgenic Arabidopsis enhanced seedling thermotolerance by decreased relative electrolyte leakage and MDA content under heat stress treatment when compared to WT plants. In addition, the induced expression of HSP101, HSFA2, and delta 1-pyrroline-5-carboxylate synthase (P5CS) under heat stress was more pronounced in transgenic plants than in WT plants. These results support the positive role of PmHSP17.9 in response to heat stress treatment.     

Binding of ferredoxin NADP+ oxidoreductase (FNR) to plant photosystem I

The binding of FNR to PSI has been postulated long ago, however, a clear evidence is still missing. In this work, using isothermal titration calorimetry (ITC), we found that FNR binds to photosystem I with its light harvesting complex I (PSI-LHCI) from C. reinhardtii with a 1:1 stoichiometry, a Kd of ~0.8 μM and ?H of ?20.7 kcal/mol. Titrations at different temperatures were used to determine the heat capacity change, ?CP, of the binding, through which the size of the interface area between the proteins was assessed as ~3000 Ų. In a different set of ITC experiments, introduction of various sucrose concentrations was used to estimate that ~95 water molecules are released to the solvent. These observations support the notion of a binding site shared by few of the photosystem I - light harvesting complex I (PSI-LHCI) subunits in addition to PsaE. Based on these results, a hypothetical model was built for the binding site of FNR at PSI, using known crystallographic structures of: cyanobacterial PSI in complex with ferredoxin (Fd), plant PSI-LHCI and Fd:FNR complex from cyanobacteria. FNR binding site location is proposed to be at the foot of the stromal ridge and above the inner LHCI belt. It is expected to form contacts with PsaE, PsaB, PsaF and at least one of the LHCI. In addition, a ~4.5-fold increased affinity between FNR and PSI-LHCI under crowded 1 M sucrose environment led us to conclude that in C. reinhardtii FNR also functions as a subunit of PSI-LHCI.     

Photosynthetic responses of a wheat (Asakaze)–barley (Manas) 7H addition line to salt stress

The photosynthetic responses to salt stress were examined in a wheat (Triticum aestivum L. cv. Asakaze)–barley (Hordeum vulgare L. cv. Manas) 7H addition line having elevated salt tolerance and compared to the parental wheat genotype. For this purpose, increasing NaCl concentrations up to 300 mM were applied and followed by a 7-day recovery period. Up to moderate salt stress (200 mM NaCl), forcible stomatal closure, parallel with a reduction in the net assimilation rate (P _N), was only observed in wheat, but not in the 7H addition line or barley. Since the photosynthetic electron transport processes of wheat were not affected by NaCl, the impairment in P _N could largely be accounted for the salt-induced decline in stomatal conductance (g _s), accompanied by depressed intercellular CO₂ concentration and carboxylation efficiency. Both, P _N and nonstomatal limitation factors (L_ns) were practically unaffected by moderate salt stress in barley and in the 7H addition line due to the sustained g _s, which might be an efficient strategy to maintain the efficient photosynthetic activity and biomass production. At 300 mM NaCl, both P _N and g _s decreased significantly in all the genotypes, but the changes in P _N and L_ns in the 7H addition line were more favourable similar to those in wheat. The downregulation of photosynthetic electron transport processes around PSII, accompanied by increases in the quantum yield of regulated energy dissipation and of the donor side limitation of PSI without damage to PSII, was observed in the addition line and barley during severe stress. Incomplete recovery of P _N was observed in the 7H addition line as a result of declined PSII activity probably caused by enhanced cyclic electron flow around PSI. These results suggest that the better photosynthetic tolerance to moderate salt stress of barley can be manifested in the 7H addition line which may be a suitable candidate for improving salt tolerance of wheat.