A DESD-box helicase functions in salinity stress tolerance by improving photosynthesis and antioxidant machinery in rice (Oryza sativa L. cv. PB1)

The exact mechanism of helicase-mediated salinity tolerance is not yet understood. We have isolated a DESD-box containing cDNA from Pisum sativum (Pea) and named it as PDH45. It is a unique member of DEAD-box helicase family; containing DESD instead of DEAD/H. PDH45 overexpression driven by constitutive cauliflower mosaic virus-35S promoter in rice transgenic [Oryza sativa L. cv. Pusa Basmati 1 (PB1)] plants confers salinity tolerance by improving the photosynthesis and antioxidant machinery. The Na⁺ ion concentration and oxidative stress parameters in leaves of the NaCl (0, 100 or 200 mM) treated PDH45 overexpressing T₁ transgenic lines were lower as compared to wild type (WT) rice plants under similar conditions. The 200 mM NaCl significantly reduced the leaf area, plant dry mass, net photosynthetic rate (P_N), stomatal conductance (gs), intercellular CO₂ (Ci), chlorophyll (Chl) content in WT plants as compared to the transgenics. The T₁ transgenics exhibited higher glutathione (GSH) and ascorbate (AsA) contents under salinity stress. The activities of antioxidant enzymes viz. superoxide dismutase (SOD), ascorbate peroxidase (APX), guaiacol peroxidase (GPX) and glutathione reductase (GR) were significantly higher in transgenics; suggesting the existence of an efficient antioxidant defence system to cope with salinity induced-oxidative damage. Yeast two-hybrid assay indicated that the PDH45 protein interacts with Cu/Zn SOD, adenosine-5′-phosphosulfate-kinase, cysteine proteinase and eIF(4G), thus confirming the involvement of ROS scavenging machinery in the transgenic plants to provide salt tolerance. Furthermore, the T₂ transgenics were also able to grow, flower, and set viable seeds under continuous salinity stress of 200 mM NaCl. This study provides insights into the mechanism of PDH45 mediated salinity stress tolerance by controlling the generation of stress induced reactive oxygen species (ROS) and also by protecting the photosynthetic machinery through a strengthened antioxidant system.     

Suppression of tomato <Emphasis Type="Italic">SlGGP</Emphasis> aggravates methyl viologen-mediated oxidative stress

Ascorbate (AsA) is an important antioxidant that can scavenge reactive oxygen species to protect plant cells against oxidative stress. Guanosine 5'-diphosphate (GDP)-L-galactose phosphorylase (GGP) is a key enzyme in the AsA biosynthetic pathway. To investigate the functions of GGP in AsA synthesis and oxidative stress tolerance in tomato, antisense lines with a reduced expression of SlGGP were obtained. Photobleaching after treatment of leaf disks with methyl viologen was more severe in transgenic lines compared to wild type (WT) plants. Moreover, compared with the WT plants, the transgenic plants showed a higher content of hydrogen peroxide, superoxide anion, malondialdehyde, as well as ion leakage, but a lower content of AsA and chlorophylls, ascorbate peroxidase activity, net photosynthetic rate, and maximal photochemical efficiency of photosystem II. Results of real-time quantitative polymerase chain reaction show that suppression of the SlGGP gene in the transgenic plants reduced their oxidative stress tolerance.     

A novel stress-associated protein SbSAP14 from Sorghum bicolor confers tolerance to salt stress in transgenic rice

We have characterized a member of the stress-associated protein (SAP) gene family from Sorghum bicolor (SbSAP14) with A20 and AN1 zinc-finger domains. Expression profiling revealed that SbSAP14 is specifically induced in response to dehydration, salt, and oxidative stress as well as abscisic acid treatment. During the early stage of salt stress, overexpression of SbSAP14 was able to prevent yellowing and withering of the leaf tip of rice plants. Measurements of malondialdehyde, ion leakage, and chlorophyll content demonstrated that transgenic rice had an enhanced tolerance to oxidative damage caused by salt stress. Under prolonged salt stress, transgenic rice plants had a higher seed germination rate and higher percentage seedling survival than wild-type (WT) plants. Importantly, in vivo and in situ assays revealed that the accumulation of reactive oxygen species in transgenic rice plants was significantly lower than that in WT plants. Among the six antioxidant genes tested, APX2, CatB, CatC, and SodA1 showed a higher expression level in transgenic rice than in WT rice. Based on these results, we propose that SbSAP14 may play a key role in antioxidant defense systems and possibly be involved in the induction of antioxidant genes in plants, suggesting a possible mechanism of the SAP gene family in stress defense response.     

Altered gibberellin content affects growth and development in transgenic tobacco lines overexpressing a wheat gene encoding F-box protein

In a previous study, we have identified and characterized gene from wheat (Triticum aestivum L.) encoding F-box protein and named it TaFBA. In this paper, transgenic tobacco (Nicotiana tabacum L.) plants overexpressing TaFBA1 displayed accelerated growth early, but the rate slowed gradually at later stages of growth, and the mature transgenic plants were even shorter in stature and flowered later than did the wild type (WT). Treatment with gibberellin (GA) conferred an accelerated growth rate to the transgenic tobacco plants at later stages, similar to that of WT, whereas growth was inhibited more seriously in WT than in transgenic tobacco when plants were treated with a GA biosynthesis inhibitor. The content of GA in transgenic tobacco plants was higher at early developmental stages, but it was lower at later growth stages than in WT. Some GA biosynthesis genes were down regulated, which was accompanied with elevated expression of a GA catabolism gene. Thus, our results suggest that TaFBA1 is possibly involved in the regulation of plant growth and development, and that it may be related to the production, metabolism, and proper function of GA.     

Cloning and Functional Characterization of a Novel Glutathione <Emphasis Type="Italic">S</Emphasis>-Transferase Gene from <Emphasis Type="Italic">Limonium bicolor</Emphasis>

Plant glutathione S-transferases (GSTs) are involved in protecting plants against both diverse biotic and abiotic stresses. In the present study, a novel GST gene (LbGST1) was cloned from Limonium bicolor (Bunge) Kuntze (Plumbaginaceae). To characterize its function in salt tolerance, tobacco lines transformed with LbGST1 were generated. Compared with wild-type (WT) tobacco, transgenic plants overexpressing LbGST1 exhibited both GST and glutathione peroxidase activities. Moreover, superoxide dismutase, peroxidase (POD), and catalase activities in transgenic plants were significantly higher than those in WT plants, particularly when grown under conditions of salt stress. Similarly, levels of proline in transgenic plants were also higher than those in WT plants grown under NaCl stress conditions. Whereas, Malondialdehyde contents in transgenic plants were lower than those in WT plants under NaCl conditions. Furthermore, Na⁺ content in transgenic plants was lower than that in WT plants under these stress conditions. Subcellular localization analysis revealed that the LbGST1 protein was localized in the nucleus. These results suggested that overexpression of LbGST1 gene can affect many physiological processes associated with plant salt tolerance. Therefore, we hypothesize that LbGST1 gene can mediate many physiological pathways that enhance stress resistance in plants.     

The involvement of wheat U‐box E3 ubiquitin ligase TaPUB1 in salt stress tolerance

U‐box E3 ubiquitin ligases play important roles in the ubiquitin/26S proteasome machinery and in abiotic stress responses. TaPUB1‐overexpressing wheat (Triticum aestivum L.) were generated to evaluate its function in salt tolerance. These plants had more salt stress tolerance during seedling and flowering stages, whereas the TaPUB1‐RNA interference (RNAi)‐mediated knock‐down transgenic wheat showed more salt stress sensitivity than the wild type (WT). TaPUB1 overexpression upregulated the expression of genes related to ion channels and increased the net root Na⁺ efflux, but decreased the net K⁺ efflux and H⁺ influx, thereby maintaining a low cytosolic Na⁺/K⁺ ratio, compared with the WT. However, RNAi‐mediated knock‐down plants showed the opposite response to salt stress. TaPUB1 could induce the expression of some genes that improved the antioxidant capacity of plants under salt stress. TaPUB1 also interacted with TaMP (Triticum aestivum α‐mannosidase protein), a regulator playing an important role in salt response in yeast and in plants. Thus, low cytosolic Na⁺/K⁺ ratios and better antioxidant enzyme activities could be maintained in wheat with overexpression of TaPUB1 under salt stress. Therefore, we conclude that the U‐box E3 ubiquitin ligase TaPUB1 positively regulates salt stress tolerance in wheat.     

Overexpression of a peroxiredoxin gene from <Emphasis Type="Italic">Tamarix hispida</Emphasis>, <Emphasis Type="Italic">ThPrx1</Emphasis>, confers tolerance to oxidative stress in yeast and Arabidopsis

Peroxiredoxins (Prxs) are ubiquitous thiol-specific antioxidant enzymes that are critically involved in cell defense and protect cells from oxidative damage. In this study, a putative Type II Prx (ThPrx1) was identified and characterized from Tamarix hispida. The expression of ThPrx1 is highly induced in response to hydrogen peroxide (H₂O₂) and methyl viologen (MV) stresses. When expressed ectopically, ThPrx1 showed enhanced tolerance against oxidative stress in yeast and Arabidopsis. In addition, transgenic Arabidopsis plants overexpressing ThPrx1 displayed improved seedling survival rates and increased root growth and fresh weight gain under H₂O₂ and MV treatments. Moreover, transgenic Arabidopsis plants showed decreased accumulation of H₂O₂, superoxide (O₂^??) and malondialdehyde (MDA), increased superoxide dismutase (SOD) activity compared to wild-type (WT) plants under oxidative stress. Moreover, transgenic plants maintained higher photosynthesis efficiency and lower electrolyte leakage rates than that of WT plants under stress conditions. These results clearly indicated that ThPrx1 plays an important role in cellular redox homeostasis under stress conditions, leading to the maintenance of membrane integrity and increased tolerance to oxidative stress.     

Overexpression of Pp14-3-3 from Pyrus pyrifolia fruit increases drought and salt tolerance in transgenic tobacco plant

Drought and salinity are the major abiotic stresses, which reduce agricultural productivity. In plants, 14-3-3s function as regulators of many target proteins through direct protein-protein interactions and play an important role during response to abiotic stresses. Here we report that CaMV 35S promoter driven overexpression of a Pyrus pyrifolia 14-3-3 gene, Pp14-3-3, improves drought and NaCl tolerance in T1 generation plants of transgenic tobacco (Nicotiana tabacum L. cv Xanthi). Under drought and NaCl stresses, the Pp14-3-3 was largely expressed in T1 transgenic tobacco lines, and compared with the wild-type (WT), transgenic tobacco plants showed relatively normal growth condition. In addition, the levels of membrane lipid peroxidation in T1 transgenic lines were definitely lower than that in WT according to the significantly decreased content of malondialdehyde. Meanwhile, the T1 transgenic tobacco lines showed significantly slower superoxide anion production rate than the WT under abiotic stress. Moreover, both the glutathione S-transferase (GST) and ascorbate peroxidase (APX) activities in T1 transgenic lines were markedly higher than those in WT. GSTs and APXs are important components of plant antioxidant system, and the results of present study suggested that Pp14-3-3 should play a crucial role in reducing oxidative damage caused by drought and salt stresses.     

Overexpression of Loquat Dehydrin Gene <Emphasis Type="Italic">EjDHN1</Emphasis> Promotes Cold Tolerance in Transgenic Tobacco

Dehydrins (DHNs) play vital roles in response to dehydration stress in plants. To examine the contribution of EjDHN to low-temperature stress in loquat (Eriobotrya japonica Lindl.), EjDHN1 was overexpressed in tobacco (Nicotiana tabacum L.). The plant growth of transgenic lines was significantly better than wild type (WT) after 4 d of recovery from cold stress. Cold stress led to membrane lipid peroxidation and reduced photosystem II (PSII) activity in leaves, and these were less severe in transgenic lines. To examine oxidative stress tolerance, the plants were treated with different concentrations of methyl viologen (MV), which inhibited plant growth both in WT and transgenic lines. After exposure to 2.0 μM MV for 10 d, the WT plants had a dramatically lower survival rate. MV treatment in leaf disks confirmed that transgenic lines accumulated less reactive oxygen species (ROS) and suffered less lipid peroxidation. The results suggested that the tolerance of the transgenic plants to cold was increased, and EjDHN1 could protect cells against oxidative damage caused by ROS production under cold stress. It also provided evidences that the enhanced cold tolerance resulted from EjDHN1 overexpression could be partly due to their protective effect on membranes by alleviating oxidative stresses.     

Overexpression of Rice CBS Domain Containing Protein Improves Salinity, Oxidative, and Heavy Metal Tolerance in Transgenic Tobacco

We have recently identified and classified a cystathionine ??-synthase domain containing protein family in Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa L.). Based on the microarray and MPSS data, we have suggested their involvement in stress tolerance. In this study, we have characterized a rice protein of unknown function, OsCBSX4. This gene was found to be upregulated under high salinity, heavy metal, and oxidative stresses at seedling stage. Transgenic tobacco plants overexpressing OsCBSX4 exhibited improved tolerance toward salinity, heavy metal, and oxidative stress. This enhanced stress tolerance in transgenic plants could directly be correlated with higher accumulation of OsCBSX4 protein. Transgenic plants could grow and set seeds under continuous presence of 150?mM NaCl. The total seed yield in WT plants was reduced by 80%, while in transgenic plants, it was reduced only by 15?C17%. The transgenic plants accumulated less Na⁺, especially in seeds and maintained higher net photosynthesis rate and Fv/Fm than WT plants under NaCl stress. Transgenic seedlings also accumulated significantly less H₂O₂ as compared to WT under salinity, heavy metal, and oxidative stress. OsCBSX4 overexpressing transgenic plants exhibit higher abiotic stress tolerance than WT plants suggesting its role in abiotic stress tolerance in plants.     

过表达ZmSKIP基因提高烟草抗低温胁迫的能力

以野生型和过表达ZmSKIP基因烟草为试材, 研究了低温胁迫下过表达ZmSKIP对烟草抗氧化能力的影响。测定了不同低温处理时间下过表达ZmSKIP转基因烟草T3代植株和野生型植株抗氧化酶如超氧化物歧化酶（SOD）、过氧化氢酶（CAT）、过氧化物酶（POD）活性和丙二醛（MDA）含量以及相对电导率, 结果表明, 低温下, 相对于野生型植株, 转基因烟草具有较高的抗氧化酶活性和较低的相对电导率和MDA含量, 说明过表达ZmSKIP提高了转基因植株的耐低温胁迫能力。     

Effect of water stress on yield and nutrition quality of tomato plant overexpressing StAPX

We investigated the effect of water stress on yield and quality of tomato plants overexpressing Solanum lycopersicum thylakoid-bound ascorbate peroxidase gene (StAPX). APX activity, hydrogen peroxide content, net photosynthetic rate of tomato leaves, and yield and nutrition quality of tomato fruits were measured under soil moisture 70, 60, and 50 % of full field capacity. Results show that the capability of APX for scavenging hydrogen peroxide induced by water stress was higher in the transgenic than the wild type (WT) plants. The yield of fruits of the transgenic tomato plants was higher than that of WT plants under water stress and the fruit nutrition quality was not different. These results indicate that overexpression of StAPX might improve water stress tolerance in the transgenic tomato plants.     

The garlic NF-YC gene, <Emphasis Type="Italic">AsNF-YC8</Emphasis>, positively regulates non-ionic hyperosmotic stress tolerance in tobacco

To investigate the relationship between nuclear factor Y (NF-Y) and stress tolerance in garlic, we cloned a NF-Y family gene AsNF-YC8 from garlic, which was largely upregulated at dehydrate stage. Expression pattern analyses in garlic revealed that AsNF-YC8 is induced through abscisic acid (ABA) and abiotic stresses, such as NaCl and PEG. Compared with wild-type plants, the overexpressing-AsNF-YC8 transgenic tobacco plants showed higher seed germination rates, longer root length and better plant growth under salt and drought stresses. Under drought stress, the transgenic plants maintained higher relative water content (RWC), net photosynthesis, lower levels of malondialdehyde (MDA), and less ion leakage (IL) than wild-type control plants. These results indicate the high tolerance of the transgenic plants to drought stress compared to the WT. The transgenic tobacco lines accumulated less reactive oxygen species (ROS) and exhibited higher antioxidative enzyme activities compared with wild-type (WT) plants under drought stress, which suggested that the overexpression of AsNF-YC8 improves the antioxidant defense system by regulating the activities of these antioxidant enzymes, which in turn protect transgenic lines against drought stress. These results suggest that AsNF-YC8 plays an important role in tolerance to drought and salt stresses.