Proline is emerging as a critical component of drought tolerance and fine tuning of its metabolism under stress affects the plants sensitivity and response to stress. Thus the study was carried out to analyse the effect of water deficit on the proline content and principal enzymes involved in its synthesis (Δ¹-pyrolline-carboxylate synthetase) and catabolism (proline dehydrogenase) at different developmental stages and in different organs (roots, nodules, leaves, pod wall, and seeds) of two chickpea (Cicer arietinum L.) cultivars differing in drought tolerance (drought tolerant ICC4958 and drought sensitive ILC3279). It was observed that increased Δ¹-pyrolline-carboxylate synthetase activity under moderate stress in roots and nodules of ICC4958 caused an increase in proline content during initiation of reproductive development whereas increased proline dehydrogenase activity in nodules and leaves at this period helped to maintain reducing power and energy supply in tissues and proper seed development as seed biomass increased consistently up to maturity. On the other hand, roots and nodules of ILC3279 responded to stress by increasing proline content after the developmental phase of reproductive organs was over (near maturity) which negatively affected the response of pod wall to stress. Concurrent increase in activities of Δ¹-pyrolline-carboxylate synthetase and proline dehydrogenase in pod wall of ILC3279 aggravated the oxidative stress and affected seed development as seed biomass initially increased rapidly under stress but was unaffected near maturity.

Rice (Oryza sativa L.) is a salt-sensitive species. Salt stress can cause injury to the plant cellular membrane. Plant lipid transfer proteins (LTPs) are abundant lipid binding proteins that are important in membrane vesicle biogenesis and trafficking, however, the biological importance of LTPs on salt-stress response in rice remains unclear. Therefore, salt-responsive rice LTPs were identified and characterized in this study. Microarray analysis showed seven genes positively regulated by salinity, including five Ltp genes (LtpII.3, LtpII.5, LtpII.6, LtpV.1, and LtpV.2) and two Ltp-like (LtpL; LtpL1, and LtpL2) genes. Amino acid alignment revealed that all these Ltp and LtpL genes contained the N-terminal signal peptide. Apart from LtpL1, all salt-inducible Ltp genes had the conserved eight cysteine residue motifs backbone. Verification of gene expression to different stimuli in rice seedlings revealed that salt-regulated Ltp genes differentially responded to drought, cold, H₂O₂, abscisic acid (ABA) and CaCl₂. Furthermore, the expression of Ltp and LtpL genes was tissue-specifically regulated by ABA-dependent and independent pathway. In silico analysis of a 1.5-kb 5'-upstream region of these genes showed regulatory cis-elements associated with ABA, calcium, and cold/drought responses. Three LtpII subfamily genes, including LtpII.3, LtpII.5, and LtpII.6, were strictly expressed in flowers and seeds, and LtpIII.1 mRNA strongly accumulated in stem tissue. Subcellular localization analysis of LTP-DsRed fusion proteins revealed that the five LTPs and two LTPLs localized at the endoplasmic reticulum. The results provide new clues to further understanding the biological functions of Ltp genes.

The tonoplast and plasma membrane localized sodium (potassium)/proton antiporters have been shown to play an important role in plant resistance to salt stress. In this study, AtNHX1 and AtNHX3, two tonoplast Na⁺(K⁺)/H⁺ antiporter encoding genes from Arabidopsis thaliana, were expressed in poplar to investigate their biological functions in the resistance to abiotic stresses in woody plants. Transgenic poplar plants expressing either gene exhibited increased resistance to both salt and water-deficit stresses. Compared to the wild type (WT) plants, transgenic plants accumulated more sodium and potassium ions in the presence of 100 mM NaCl and showed reduced electrolyte leakage in the leaves under water stress. Furthermore, the proton-translocating and cation-dependent H⁺ (Na⁺/H⁺ or K⁺/H⁺) exchange activities in the tonoplast vesicles isolated from the leaves of transgenic plants were higher than in those isolated from WT plants. Therefore, constitutive expression of either AtNHX1 or AtNHX3 genetically modified the salt and water stress tolerance of transgenic poplar plants, providing a potential tool for engineering tree species with enhanced resistance to multiple abitotic stresses.

The relationship between hydrogen peroxide (H₂O₂) and nitric oxide (NO) in the regulation of ascorbate and glutathione metabolism by jasmonic acid (JA) in Agropyron cristatum leaves were studied. Results showed that JA increased the production of H₂O₂ and NO, the activities of ascorbate peroxidase (APX), glutathione reductase (GR), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), L-galactono-1,4-lactone dehydrogenase (GalLDH), and γ-glutamylcysteine synthetase (γ-ECS), as well as transcription of the respective genes and also the content of reduced ascorbate (AsA) and reduced glutathione (GSH). Above increases were suppressed by pre-treatments with H₂O₂ synthesis inhibitor diphenylene iodonium (DPI), H₂O₂ scavenger dimethylthiourea (DMTU), NO synthesis inhibitor N ^G-nitro-L-Arg methyl ester (L-NAME), and NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO). Pre-treatments with DPI and DMTU reduced H₂O₂ and NO production. Pre-treatments with L-NAME and cPTIO reduced NO production, but did not reduce the H₂O₂ production induced by JA. Our results suggested that NO acted downstream of H₂O₂ in JA signalling in the up-regulation of ascorbate and glutathione metabolism in A. cristatum leaves.

Kinetin-induced programmed cell death of field bean (Vicia faba spp. minor) root cortex cells led to aerenchyma formation. The process was accompanied by appearance of a greater amount of reactive oxygen species (ROS), greater superoxide dismutase (SOD) and catalase (CAT) activities, as well as by thickening cell walls and changes in sugar amounts, particularly in cell wall-bound sugars. The obtained results justify the supposition that ROS scavengers together with an increased amount of sugars (soluble, storage, and cell wall-bound) and thick cell walls protected the cells against death. Thus, kinetin played a dual role because it induced programmed death of chosen cells and simultaneously stimulated protective mechanisms against death in other cells. These results confirm an earlier suggestion that cell death induced by kinetin is a specific process during which its progression is hallmarked by metabolic and morphological features.

Litchi is one of the most important subtropical evergreen fruit trees in Southern Asia. Litchi floral buds are a mix of axillary or apical panicle primordia, leaf primordia, and rudimentary leaves. Under usual winter and early spring conditions, the axillary panicle primordia prevail, and the rudimentary leaves abscise when low temperatures reach a certain threshold. The floral buds ultimately develop into pure panicles. Understanding the regulatory mechanism of rudimentary leaf senescence is of great importance for litchi flowering. In this study, litchi potted trees at the floral differentiation stage were treated with low and high temperatures in order to induce senescence or development of leaves. The microstructure of the petiole base of the rudimentary leaves was determined. The results show several layers of flattened cells forming in the abscission zone of the rudimentary leaves that were treated with low temperatures as well as an obvious boundary regarded as the abscission layer zone. We also determined the gene expression in the leaves with different developmental fate. The results show that the LcRboh, LcMC-1-like, and LcPirin genes were significantly induced in the rudimentary leaves treated with low temperatures, and the expression increased with the proceeding of senescence. The expression of the genes encoding class Ι β-1,3-glucanase and β-xylosidase also increased with the senescence, suggesting their possible involvement in the low temperature-induced senescence of the rudimentary leaves.

The root specificity and phosphate (Pi) deficiency responsiveness of high-affinity phosphate transporter (PHT1) genes point to their promoters as a sustainable system to drive Pi acquisition-related transgenes in plants. In this study, a 3-kb promoter of the AtPHT1;4 gene from Arabidopsis thaliana fused to the β-glucuronidase (GUS) reporter gene was biolistically introduced into wheat (Triticum aestivum L.) and functionally characterized in transgenic plants grown in hydroponics and in pots with soil under various Pi supply rates. From among 27 T₁ progeny derived from 250 T₀, four transgenic lines reached T₃, with two of them showing detectable GUS activity in the roots of T₄ plants. An unusually high number of transgene insertions characterized these transgenic lines, along with an irregular pattern of histochemical GUS staining and weak GUS activity. GUS expression driven by AtPHT1;4 was consistently higher under most assay conditions, as it was unaffected by 0 to 0.5 mM Pi in hydroponically grown plants, as well as by 16 to 20 mg(P) kg^-1(soil) in potted plants. Raising the soil P up to or above 40 mg kg^-1 significantly down-regulated the quantity of GUS transcripts. These results show that the responsiveness of the AtPHT1;4 promoter to Pi availability in transgenic wheat was restricted to soil-grown plants, which highlighted the relevance of the substrate and Pi supply rates in assessing molecular responses to Pi deficiency.

Physiological responses of Oryza sativa L. to lead excess (10 and 50 μM) were studied in a hydroponic system after 48- and 96-h exposure. Accumulation of Pb in stressed rice shoots was concomitant with an increased metal concentration in the growth media and duration of exposure. The Pb stress resulted in an enhanced lipid peroxidation accompanied by altered activities of antioxidants. A substantial increase in α-tocopherol content of the Pb stressed rice shoots was observed suggesting its important role as an antioxidant. Among the antioxidant enzymes studied, activities of superoxide dismutase (SOD) and ascorbate peroxidase (APX) increased in the Pb-treated rice shoots, whereas that of catalase (CAT) declined. Activity of an important ascorbate-glutathione cycle enzyme, glutathione reductase (GR), also increased significantly in the Pb-treated shoots. The results suggest that Pb toxicity resulted in induction of oxidative stress in rice shoots, and α-tocopherol accumulation and upregulation of SOD, APX, and GR activities play an effective role in acclimatization to Pb stress.

The mechanisms of growth inhibition and antioxidative response were investigated in wheat roots exposed to 300 μM iron together with different zinc concentrations (0, 50, and 250 μM). All Zn concentrations decreased Fe content but increased Zn content in the roots and leaves of Fe-treated seedlings. Compared with Fe stress alone, 50 or 250 μM Zn + Fe treatment stimulated root growth, and increased cell viability but decreased malondialdehyde content, which were correlated with the decreases of total and apoplastic hydrogen peroxide and superoxide anion radical (O₂ ^.-) content along with apoplastic hydroxyl radical content. Generation of O₂ ^.- in response to 10 μM diphenylene iodonium suggested that NADPH oxidase activity was lower in Zn + Fe-treated roots than in other roots. In addition, cell wallbound peroxidase, diamine oxidase, and polyamine oxidase in Fe-treated roots were insensitive to Zn addition. Further study showed the stimulation of total superoxide dismutase and glutathione reductase (GR) activities as well as apoplastic catalase, ascorbate peroxidase, and GR in Zn + Fe-stressed roots in comparison with Fe-alone-treated ones. Taken together, Zn could alleviate iron-inhibitory effect on root growth, which might be associated with the decrease of lipid peroxidation, the increase of cell viability and the reductions of reactive oxygen species generation.

Within the last two decades, plant viral vectors have emerged as an excellent tool for the expression of foreign peptides and proteins. Virus particles carrying foreign antigenic epitopes present some interesting advantages for vaccine design and other applications. This review covers recent advances in the use of some typical plant viruses with helical particles that present heterologous peptides with particular emphasis on particles derived from the Potato virus X (PVX) and its uses.

Zea mays L. is less tolerant to drought than Sorghum bicolor L. In the present study, we investigated the response of both plants to drought stress applied under field conditions by withholding water for 10 d. The plant growth in terms of shoot fresh and dry masses was more severely reduced in maize than in sorghum, consistently with reduction of leaf relative water content. Gas exchange was also more inhibited by drought in maize than in sorghum. The water use efficiency (WUE) of maize fluctuated during the day and in response to the drought stress. In contrast, sorghum was able to maintain a largely constant WUE during the day in the well-watered plants as well as in the stressed ones. Studying the expression of four aquaporin genes (PIP1;5, PIP1;6, PIP2;3, and TIP1;2) revealed that PIP1;5 in leaves and PIP2;3 in roots were highly responsive to drought in sorghum but not in maize, where they might have supported a greater water transport. The expression pattern of PIP1;6 suggests its possible role in CO₂ transport in control but not droughty leaves of both the plants. TIP1;2 seemed to contribute to water transport in leaves of the control but not droughty plants. We conclude that PIP1;5 and PIP2;3 may have a prominent role in drought tolerance and maintenance of WUE in sorghum plants.

As key nuclear transcription factors, the ethylene-insensitive3/EIN3-like (EIN3/EIL) proteins play important roles in ethylene signal transduction pathway in various plants. In order to better understand the role of EIN3/EILs, one EIN3-like gene (designated ZmEIL1) was isolated from maize (Zea mays L.). The full-length cDNA of ZmEIL1 was 1 999 bp in length and encoded 647 amino acids. Sequence comparison of ZmEIL1 protein with other EIN3/EILs proteins revealed high conservation of five α-helices that could form a V-shaped cleft in a 3-D model, just like AtEIL3 in Arabidopsis thaliana. This protein showed transcriptional activation and activation domain located on the 507 - 647 amino acids in yeast. Furthermore, ZmEIL1 could interact with ZmERF1 in the yeast systems, which was downstream response factor in ethylene signal transduction pathway. Its mRNA could be highly induced in maize seedlings by ethephon and 1-methylcyclopropene treatments. Meanwhile, ZmEIL1 showed relatively high expression at 20 d after pollination in maize kernel. These results show that ZmEIL1 played an important role in the growth and development by participating in ethylene signalling pathway in maize.

Providing sufficient food to burgeoning population from the steadily shrinking arable land seems to be very difficult in near future and is one of the foremost challenges for plant scientists. In addition, there are several biotic and abiotic stresses which frequently encounter crop plants during various stages of life cycle, resulting in considerable yield losses. Environmental stresses, including drought, flooding, salinity, temperature (both low and high), high radiation, and xenobiotics induce toxicity, membrane damage, excessive reactive oxygen species (ROS) production, reduced photosynthesis, and altered nutrient acquisition. Several indigenous defence mechanisms (physiological and molecular) are triggered in plants on exposure to environmental cues. Enhancement of resistance of crop plants to environmental stresses has been the topic of prime interest for agriculturalists and plant scientists since long. Development of water and salinity stress-tolerant crops through genetic engineering provides an avenue towards the reclamation of farmlands that have been lost due to salinity and lack of irrigation water/rainfall. Understanding the complexity of stress tolerance mechanisms in orthodox or model plants at the genetic and molecular levels improves feasibility of enhancing tolerance of sensitive crop plants.

Drought and salinity are severe environmental stresses and limit soybean growth. In this study, a comparative analysis of physiological and molecular responses of two soybean (Glycine max L.) genotypes to these stresses was carried out. Plants of drought-tolerant genotype RD (cv. FD92) and sensitive genotype SD (cv. Z1303) were exposed to 15 % (m/v) PEG 6000, which simulated drought stress, or 150 mM NaCl. The RD plants maintained larger leaf area and higher net photosynthetic rate, chlorophyll content, stomatal conductance, and relative water content compared with the SD plants. Leaf proline content increased under both stresses more in RD than in SD. The drought tolerance of RD plants was also correlated with greater antioxidant activity and lower content of hydrogen peroxide and malondialdehyde under stress conditions. Amounts of abscisic acid, jasmonic acid, and salicylic acid under stress increased to a greater extent in RD than in SD plants. At the molecular level, the effects of 20-d stress treatments were manifested by relatively higher expression of drought- or salt-related genes: GmP5CS, GmDREB1a, GmGOLS, GmBADH, and GmNCED1 in RD plants than in SD plants. These results form the basis for understanding the mechanisms of the drought- and salt-stress tolerance in soybean.

Salt stress is one of the abiotic stresses limiting the yield of crops worldwide. However, the molecular mechanisms underlying the regulation of plant response to salt stress are not completely elucidated. Ethylene response factors (ERFs) are a subfamily of the AP2 (APETALA2)/ERF transcription factor family that regulates multiple aspects of plant growth and development, and plant responses to biotic and abiotic stresses. ERF96 is one of the small ERFs that is involved in plant defense response and abscisic acid signaling in Arabidopsis. By using real time quantitative PCR, we found that the expression of ERF96 in the wild type Arabidopsis thaliana (cv. Col-0) seedlings was induced by NaCl treatment. The transgenic plants overexpressing ERF96 were more tolerant to salt stress in terms of NaCl inhibited seed germination, early seedling development, and fresh mass. Consistent with these observations, elevated expressions of some NaCl-responsive genes including responsive drought 29 (RD29A), Δ¹-pyrroline-5-carboxylate synthetase (P5CS), cold responsive 15A (COR15A), and kinase 1 (KIN1) were observed in the transgenic plants in the presence of NaCl. We also found that the Na⁺ and K⁺ content and expressions of genes related to Na⁺/K⁺ homeostasis including stelar K⁺ outward rectifier (SKOR) and potassium transport 2/3 (AKT2/3) were altered in the ERF96 transgenic plants in response to NaCl treatment. Taken together, these results showed that overexpression of ERF96 enhanced plant tolerance to salt stress, indicating that ERF96 is a positive regulator of salt tolerance in Arabidopsis.

Cassava (Manihot esculenta Crantz) is an important crop, and its starch formation is regulated by sucrose metabolism and transport. To understand the roles of sucrose metabolism and transport in cassava under water stress, we studied not only sucrose metabolism and transport in cassava abscission zones (AZs) but also expression of respective genes. Sucrose was transported from leaves to roots in the early stage of water stress, and a reverse sucrose flow was detected in the later stages of the stress. The decrease in sucrose content was related to leaf senescence and inhibition of photosynthesis. Microarray analyses showed seven genes encoding sucrose synthase, nine genes encoding sucrose transporters, and eight genes encoding invertase in the cassava AZs under the water stress. Reverse transcription quantitative PCR confirmed two sucrose synthase and two invertase genes significantly upregulated under the stress, whereas one sucrose transporter gene was downregulated. The sucrose synthase and invertase gene expressions were negatively correlated with sucrose content under water stress, whereas sucrose transporter gene expressions were positively correlated with sucrose content.

The present study was conducted to examine differential responses of roots and leaves of Artemisia annua to different arsenic concentrations (50, 100, and 150 μΜ) and treatment durations (1, 3, 5, or 7 d). The values of bioconcentration factor and translocation factor calculated on the basis of total As-accumulation in roots and shoots suggested that A. annua is a good As-accumulator. Above and below ground plant biomass was enhanced at 100 μΜ As but at 150 μΜ As was significantly reduced. As-treatment caused membrane damage more in the roots than in the leaves as reflected by higher degree of lipid peroxidation in the roots than in the leaves. In response to As stress, plants activated antioxidative defense for detoxification of induced reactive oxygen species (ROS), As sequestration via phytochelatins (PCS) as well as production of a wide range of secondary metabolites. All of them were activated differently in roots and leaves. Among enzymatic antioxidants, leaves significantly elevated superoxide dismutase (SOD), ascorbate peroxidase, and glutathione reductase, whereas in roots SOD, catalase, and peroxidase played significant role in ROS detoxification. Plants activated As-sequestration pathway through thiols, glutathione, and PCS and their respective genes were more induced in leaves than in roots. Further gas chromatography in tandem with mass spectroscopy analysis revealed differential modulation of secondary metabolites in leaves and roots to sustain As-stress. For example, roots synthesized linoleic acid (4.85 %) under As-treatment that probably stimulated stress-signalling pathways and in turn activated differential defense mechanisms in roots to cope up with the adverse effects of As.

During the normal developmental process, programmed gene expression is an essential phenomenon in all organisms. In eukaryotes, DNA methylation plays an important role in the regulation of gene expression. The extent of cytosine methylation polymorphism was evaluated in leaf tissues collected from the greenhouse grown plants and in in vitro-derived callus of three lowbush and one hybrid blueberry genotypes, using methylation-sensitive amplification polymorphism (MSAP) technique. Callus formation started from the leaf segments after 4 weeks of culture on a thidiazuron (TDZ) containing medium. Maximum callus formation (98 %) was observed in the hybrid blueberry at 1.0 mg dm^-3 TDZ. Although noticeable changes in cytosine methylation pattern were detected within the MSAP profiles of both leaf and callus tissues, methylation events were more polymorphic in calli than in leaf tissues. The number of methylated CCGG sites varied significantly within the genotypes ranging from 75 to 100 in leaf tissues and from 215 to 258 in callus tissues. Differences in the methylation pattern were observed not only in a tissue-specific manner but also within the genotype in a treatment specific manner. These results demonstrated the unique effect of TDZ and the tissue culture process on DNA methylation during callus development.

Dehydration responsive element binding factor (DREB) is believed to be a stress-tolerance enhancer in plants. In the present study, a cold-binding factor (CBF)/DREB homologous gene NnDREB1 (XP_010242642.1) was isolated from lotus roots using rapid amplification of cDNA ends (RACE) and reverse transcription (RT)-PCR methods. Analysis of the deduced amino acid sequence and phylogeny classified NnDREB1 into the A-1 group of the DREB1 subfamily. Expression profiling using a quantitative PCR method revealed that NnRDEB1 was significantly induced by NaCl, mannitol, and polyethylene glycol, but not by low temperature and abscisic acid. To evaluate function of NnRDEB1, Arabidopsis thaliana was transformed with the NnDREB1 gene in a binary vector construct. The transgenic plants exhibited higher resistance to drought compared with the wild-type plants in terms of survival rates, dry and fresh masses, and chlorophyll content. In addition, overexpression of NnDREB1 resulted in higher germination rates compared with the wild type plants on MS medium containing mannitol. The expressions of downstream target stressrelated genes, including cold-regulated15B (COR15B), rare cold inducible 2B (RCI2B) and repeat domain 26 (RD26), were activated in the transgenic plants. Taken together, the results suggest that NnDREB1 might be an important protein in lotus root drought tolerance.

To investigate the biochemical and physiological responses of Masson pine (Pinus massoniana Lamb.) seedlings to lead stress, needles, stems, and roots of two-year-old seedlings were treated with ²⁰⁷PbCO₃ for 33 d and then analyzed 1 and 7 d after the treatment was completed. Chlorophyll (Chl) b responded more sensitively than Chl a to needle Pb treatment, and the Chl content in the needles significantly decreased after Pb application to roots. The malondialdehyde and proline content remained almost unchanged, but superoxide dismutase and catalase activities increased on day 1 after all ways of Pb application. The reduced glutathione (GSH) content and GSH/oxidized glutathione ratio increased on day 1 after Pb application to stem or needles compared to the controls. At 7 d after the Pb application, the increase in dehydroascorbate (DHA) content and the decrease in the ascorbate (AsA)/DHA ratio implied a decreased antioxidant capacity of AsA. The results indicated that the antioxidants were sensitive to the Pb treatments and might be involved in the Masson pine tolerance to Pb stress.

Abscisic acid (ABA) regulates various plant physiological processes, especially participates in the plant responses to harsh environments. The 9-cis-epoxycarotenoid dioxygenase (NCED) is a key enzyme in ABA biosynthesis pathway. Here, a TaNCED with an 1 887-bp open reading frame was cloned from wheat, which encodes a peptide of 628 amino acids. A chloroplast transit peptide sequence was found at the N-terminus of the TaNCED protein. Multiple sequence alignments indicate that the TaNCED protein shared high similarities with other NCEDs from different species. Real-time quantitative PCR analysis shows that expression of TaNCED was strongly up-regulated by treatments with ABA, polyethylene glycol, and drought stress, and it was down-regulated during germination of the wheat seeds. Ectopic overexpression of the TaNCED gene in Arabidopsis resulted in an increase of endogenous ABA and free proline content. A lower water loss rate and stomatal conductance of leaves were found in the transgenic plants in comparison with the wild type. Subsequently, the transgenic plants displayed an enhanced tolerance to drought stress but delayed seed germination. These data provide evidence that the TaNCED might play a primary role in regulation of ABA content during water stress and seed dormancy.

Both lignin and silicon (Si) are major players in the resistance of plants to mechanical stress (MS). Focusing on the phenolic metabolism, here we studied the short-term effects of a local MS on tobacco (Nicotiana rustica L. cv. Basmas) plants with Si (+Si, 1 mM Na₂SiO₃) and without Si (‒Si) treatments in order to see how Si may modify local and systemic responses. One week after starting the Si treatment, a half of the plants were exposed to a mechanical pressure applying 980 Pa for 24 h on the upper side of the 3^rd leaf of each plant (+MS). The rest of the plants remained unstressed (‒MS). Plants were harvested 24 h and 72 h after starting the MS and the leaves directly exposed to the mechanical stress (DMS) and those indirectly exposed to the mechanical stress (IMS) from below and above the DMS leaf were analyzed for phenolic metabolism along with the corresponding leaves from‒MS plants. In the DMS leaf, the activities of polyphenol oxidase, phenylalanine ammonia lyase, and cytosolic and covalently-bound peroxidases increased by the MS, while decreased by Si. In accordance with this in the DMS leaf, the content of soluble and cell wall-bound phenolics and lignin were enhanced by the MS but decreased by Si. Interestingly, Si influenced the pattern of response to the MS depending on whether the leaves were directly treated by the MS or not. Silicon treatment augmented MS-induced lignin accumulation in the DMS leaf while rather inhibited lignin formation in the IMS leaves. These data show that Si modified MS-mediated changes in the phenolic metabolism differently in local and systemic leaves.

Proanthocyanidins (PAs) are the main products of the flavonoid biosynthetic pathway in many plants. However, their biological function during environmental stresses in plants is rarely reported. In the present study, the effects of pretreatment with PAs on the response of cucumber (Cucumis sativus L.) seedlings to high irradiance (HI), polyethylene glycol (PEG), and cold stress were investigated. The PAs pretreament alleviated stress-induced oxidative damage in plant cells and increased the activity of alternative oxidase (AOX) and content of abscisic acid (ABA). Furthermore, PAs-pretreated seedlings suffered less damage by the stress conditions, maintained higher content of chlorophyll a+b and AOX proteins in comparison with the control. Therefore, our findings suggest that PAs might contribute to plant tolerance to environmental stresses.

Real time quantitative PCR (qPCR) is widely used in gene expression analysis for its accuracy and sensitivity. Reference genes serving as endogenous controls are necessary for gene normalization. In order to select an appropriate reference gene to normalize gene expression in Casuarina equisetifolia under salt stress, 10 potential reference genes were evaluated using real time qPCR in the leaves and roots of plants grown under different NaCl concentrations and treatment durations. GeNorm, NormFinder, and BestKeeper analyses reveal that elongation factor 1-alpha (EF1α) and ubiquitin-conjugating enzyme E2 (UBC) were the most appropriate reference genes for real time qPCR under salt stress. However, β-tubulin (βTUB) and actin 7, which were widely used as reference genes in other plant species, were not always stably expressed. The combination of EF1α, UBC, uncharacterized protein 2, DNAJ homolog subfamily A member 2, and glyceraldehyde-3-phosphate dehydrogenase should be ideal reference genes for normalizing gene expression data in all samples under salt stress. It indicates the need for reference gene selection for normalizing gene expression in C. equisetifolia. In addition, the suitability of reference genes selected was confirmed by validating the expression of WRKY29-like and expansin-like B1. The results enable analysis of salt response mechanism and gene expression in C. equisetifolia.

Silicon (Si) could enhance plant tolerance to heavy metals; however, the mechanism of Si-mediated alleviation of cadmium (Cd) toxicity in Poa annua was not clear. In this study, we found that 100 μM Cd significantly inhibited the growth of Poa annua seedlings. Furthermore, Cd enhanced the H₂O₂ and malondialdehyde content. The activities of superoxide dismutase and ascorbate peroxidase were enhanced, but the catalase and peroxidase activities were reduced by Cd treatment. Cd also altered the activity and expression of glucose-6-phosphate dehydrogenase (G6PDH) in Poa annua roots. Application of Na₃PO₄, an inhibitor of G6PDH, decreased the activity of G6PDH, the expression of G6PDH, and increased the Cd toxicity, suggesting that G6PDH is involved in the regulation of oxidative stress induced by Cd. Application of 1 mM Si alleviated the inhibition of Cd on the growth of Poa annua seedlings. Si application not only led to reduced oxidative injuries but also decreased the accumulation of Cd in Poa annua seedlings under Cd stress. Furthermore, Si decreased the activity of G6PDH and the expression of G6PDH under Cd stress, which demonstrated that Si attenuates the Cd toxicity in Poa annua probably through decreasing the expression of G6PDH under Cd stress. When G6PDH was inhibited, the alleviation impact of Si on Cd stress was abolished. Taken together, these results demonstrated that the Cd tolerance in Poa annua enhanced by Si is mainly due to the decrease of Cd uptake in roots and lowering the oxidative stress induced by Cd.

A plasmid and two isocaudamer systems, namely, NotI/Bsp120I and SpeI/XbaI/NheI, were used to construct a new type of multi-gene plant transformation vector system. This system included a transformation vector containing the restriction enzyme cutting sites Bsp120I and XbaI as well as a cloning vector containing the restriction enzyme cutting sites NotI, Bsp120I, SpeI, and NheI. The open reading frame of the new target genes was connected to the transformation vector. The original restriction enzyme cutting site disappeared after connecting to the isocaudamer. The plant transformation vector p096871, which contained Bacillus thuringiensis (Bt) genes Cry1Ac and Cry3A as well as p09X6, which contained mtlD, strD, betA, nhaA, and ostAB, were constructed using this vector system. Resistant plants were obtained after tobacco was transformed by two vectors via the Agrobacterium-mediated method. Detection by PCR revealed that all exogenous genes were inserted into the genome of tobacco. Real-time fluorescence quantification PCR, reverse transcription PCR, and ELISA detections were performed on five transgenic lines transformed by two Bt genes. Cry1Ac and Cry3A were inserted into the genome with a single copy to transcribe and express Bt toxins. The proposed vector system reduced the number of operational procedures and minimized the difficulty of the experiment.

Lignin is closely related to the lodging resistance of common buckwheat (Fagopyrum esculentum Moench.). However, the characteristics of lignin synthesis related genes have not yet been reported. We investigated the lignin biosynthesis gene expression, activities of related enzymes, and accumulation of lignin monomers during branching stage, bloom stage, and milky ripe stage by real-time quantitative PCR, UVspectrophotometry, and gas chromatography-mass spectrometry in the 2^nd internode of three common buckwheat cultivars with different lodging resistance. The results showed that lignin content and the activity of phenylalanine ammonia lyase (PAL), 4-coumarate: CoA ligase (4CL), cinnamyl alcohol dehydrogenase (CAD) and peroxidase (POD) were closely related to the lodging resistance of common buckwheat. Further, we studied gene expression of cinnamate 4-hydroxylase (C4H), caffeoyl-CoA O-methyltransferase (CCoAOMT), ferulate 5-hydroxylase (F5H), cinnamoyl-CoA reductase (CCR), and caffeic acid O-methyltransferase (COMT). The lignin biosynthesis genes were divided into three classes according to their expression pattern: 1) expression firstly increasing and then descending (PAL, 4CL, CAD, C4H, CCoAOMT, F5H, and CCR), 2) expression remaining constant during maturation (C3H), and 3) expression decreasing with maturation (COMT). The present study provides preliminary insights into the expression of lignin biosynthesis genes in common buckwheat, laying a foundation for further understanding the lignin biosynthesis.

Membrane traffic mediated by a soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) complex contributes to plant growth and development. However, the functional significance of SNAREs involved in cell wall deposition and seedling development has not been sufficiently explored. In this study, we explored the roles of R-SNAREs VAMP721 (At1g04750) and VAMP722 (At2g33120) in seedling growth of Arabidopsis thaliana by histochemical staining, fluorescence labeling, and analyzing mutant phenotypes. Our results show a massive intracellular accumulation of cellulose and callose, and an abnormal deposition of callose at the expanding cell plate in vamp721vamp722 root cells compared with the wild type. Particularly, ectopic lignin accumulation was also observed in vamp721vamp722 root cells. The alteration of cell wall components was confirmed using Fourier transform infrared analysis. Plasma membrane integrity and cell viability were disturbed in the vamp721vamp722 seedling. Morphological observation shows that vamp721vamp722 mutations impaired development of roots, hypocotyl, cotyledon, and true leaf, and inhibited lateral root formation. Confocal images reveal that green fluorescent protein-tagged VAMP721 and VAMP722 showed a similar expression pattern and were expressed throughout all cells and tissues examined, including root and shoot apical meristems and cells of hypocotyls, cotyledons, and true leaves. Taken together, our results suggest that membrane traffic mediated by VAMP721 and VAMP722 is involved in seedling growth in A. thaliana.

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) serves not only as a key enzyme in glycolysis, but also as a multifunctional protein in other biological processes, especially in response to abiotic stresses in plants. Cytosolic GAPDH (GAPC) is a typical redox protein with selected catalytic cysteine, which undergoes reversible redox post-translational modifications (RPTMs) on its thiol group by reacting with hydrogen peroxide and nitric oxide related species. Moreover, the modified GAPC may interact with certain signal transmitters such as phosphatidic acid, phospholipase D, and osmotic stress-activated protein kinase. All these observations suggest that GAPC serve as a key mediator in redox signal transduction in plants. In this review, we provide an up-to-date insight into molecular mechanisms after H₂O₂- and NO-dependent oxidation of GAPC. We also discuss GAPC catalytic functions and potential functions as a modified protein by RPTMs.

Several different concentrations of α-tocopherol were applied to Carex leucochlora after plants had been treated with high salinity (0.8 % NaCl) in a greenhouse for one month. The results revealed that 0.8 mM α-tocopherol treatment showed the greatest alleviation of growth inhibition and cell membrane damage induced by salt stress. In comparison with NaCl alone, the 0.8 mM α-tocopherol application significantly decreased the content of hydrogen peroxide and the rate of superoxide radical generation, and increased the content of chlorophyll b, carotenoids, free proline, and soluble protein, but had no effect on the content of chlorophyll a and soluble sugar. These results suggest that α-tocopherol could effectively protect C. leucochlora plants from salt stress damage presumably by quenching the excessive reactive oxygen species to protect the photosynthetic pigments and by enhancing the osmotic adjustment.