Ethylene response factor (ERF) is a key transcription factor of plant ethylene signaling pathway, which plays an important role in plant response to abiotic and biotic stresses by regulating the expression of downstream genes. However, little is known about the mechanisms of the regulation of gene expression by ERF proteins. Chloroplast is an essential organelle that is important for photosynthesis and biosynthesis of many essential metabolites. There exists an interaction between chloroplasts and the nucleus. Chloroplasts can send multiple kinds of signals to regulate the nuclear gene expression known as retrograde signaling. In our study, we have analyzed the expression of the components related to plastid retrograde signaling pathway to elucidate the mechanism of tomato ethylene responsive factor 1 (TERF1) in response to drought stress. Our results showed that TERF1 can regulate different biogenic and operational retrograde signals to regulate nuclear genes expression, which can improve plant tolerance to drought stress. We also propose a new potential of TERF1 in regulating nuclear gene expression, including regulation of different phytohormone signaling pathways and gene posttranscriptional modification triggered by different retrograde signals. Our results have enriched our knowledge about the function of ERF proteins and ethylene signaling pathway.

An inhibitor of DNA methylation 5-azacytidine (5A) is a chemical analog of the nucleoside cytidine. This study investigated the influence of 5A-induced DNA hypomethylation on the accumulation of somatic DNA mutations (nucleotide substitutions, indels) in the Actin2 3′ untranslated region, nuclear internal transcribed spacer ITS1-5.8rRNA-ITS2, and the ribulose-1,5-bisphosphate carboxylase/oxygenase gene of Arabidopsis thaliana and analyzed concurrent changes in the expression of DNA methyltransferase and DNA repair genes. The 5A treatment (20 mg per 100 g of soil) decreased DNA methylation, and the detected 5A-induced demethylation was associated with the up-regulation of the DNA methyltransferase genes: chromomethylase AtCMT3, methyltransferase AtMETI, and domains rearranged methyltransferases AtDRM1 and AtDRM2. Cultivation of plants in the presence of 5A led to a considerable increase in the number of single nucleotide substitutions in the analyzed DNA regions of 5A-treated A. thaliana. The 5A treatment significantly increased the transcriptions of 7 DNA repair genes (endonuclease AtARP, DNA demethylases AtDME and AtDML2, DNA glycosylase AtMBD4, DNA damage-binding protein AtDDB1, and photolyases AtUVR2 and AtUVR3) out of the 17 analyzed genes from the base excision repair, nucleotide excision repair, and photoreactivation pathways. However, 5A decreased the transcription of DNA 3′-phosphatase AtZDP, DNA repair protein AtRad23a, mismatch repair proteins AtMsh2 and AtMsh3. It is possible that the changes in the transcription of the DNA repair genes contributed to the detected increase in the number of single nucleotide substitutions that accumulated in the 5A-treated A. thaliana. Taken together, the data indicate that there is an interaction between the processes of DNA methylation and mutation accumulation.

Several studies revealed the important contribution of calcineurin B-like (CBLs) and CBL-interacting kinase (CIPKs) genes in transmitting stress signals in plants. Taking advantage from the genome sequences of the cultivated potato Solanum tuberosum and its wild relatives S. commersonii and S. chacoense, we identified for the first time 10 CBLs and 26 CIPKs genes in each species. The CBLs and CIPKs derived from tandem duplications indicate that these gene families in potato mainly arise through amplification mechanisms. Once annotated, we compared the par excellence model of Arabidopsis thaliana with S. commersonii, the potato model species for studying cold tolerance. We found that four ScCBL proteins (ScCBL1, ScCBL4a, ScCBL4b, and ScCBL9) started with a conserved N-myristoylation motif (MGXXXS/T), which might function in membrane targeting of the CBLs-CIPKs complex. Additionally, expression analyses of S. commersonii CBL and CIPK genes based on RNAseq revealed diverse expression patterns following various abiotic and biotic stresses and in the four tissues analyzed (flowers, leaf, roots, and tubers). Data also suggest that the ScCBLs-ScCIPKs complex may be more responsive to abiotic rather than biotic stimuli. Overall, the results described in the present work will be useful for future investigations and for functional characterization of individual CBLs and CIPKs in Solanum.

Globe artichoke (Cynara cardunculus L. var. scolymus) is rich in flavonoids which contribute to its health-promoting properties. With the aim of understanding the genetic control of flavonoid accumulation in artichoke, we isolated an artichoke full-length cDNA sequence encoding flavonoid 3'-hydroxylase (F3'H), a major enzyme of the flavonoid hydroxylation pattern. In silico studies confirmed that the deduced amino acid sequence of CcF3'H is highly similar to F3'Hs isolated from other Asteraceae. The Northern blot analysis demonstrated that CcF3'H was highly expressed in leaves and in specific parts of the heads. Its expression differed slightly among artichoke cultivars. The overexpression of CcF3'H in tobacco plants led to the accumulation of flavonoids and to an increase of flower colour intensity, thus identifying CcF3'H as promising candidate for genetic engineering. CcF3'H represents the first structural gene of the flavonoid biosynthesis isolated from C. cardunculus, and its characterization sheds light on the accumulation of flavonoids.

Two types of hairpin RNA (hpRNA) constructions were designed using a chimeric gene formed from two genes, the coat protein (CP) gene or the silencing suppressor gene, from the Cucumber mosaic virus (CMV) and the Potato virus Y (PVY_N), respectively; one type generated a single hairpin structure, whereas the other formed a two-hairpin structure. Four constructs, pDCPSH (double CP gene fragments, single hairpin), pDCPDH (double CP gene fragments, double hairpins), pHC2bSH (two silencing suppressor gene fragments, single hairpin), and pHC2bDH (two silencing suppressor gene fragments, double hairpins), were individually introduced into tobacco plants. A transcript analysis demonstrates that the small interference RNA (siRNA) processing efficiency was greater with the double-hairpin construct than with the single-hairpin construct, although the expression of their target genes were similar. A viral resistance assay shows that the transgenic tobacco plants effectively resisted a mixed infection of CMV and Potato virus Y (PVY^N) and that pDCPDH exhibited the highest silencing efficiency. The accumulation of siRNA in the inoculated transgenic plants expressing different hairpin structures was similar. A genetic analysis reveals that viral resistance in the transgenic plants was stably inherited from the T₀ to T₁ generation. A transcript analysis and a viral resistance assay indicate that the double-hairpin structure of the same target sequences tended to produce more siRNA before the virus inoculation and thus strengthened RNA-mediated viral resistance.

The objective of this study was to relate the activation of enzymatic antioxidant system to the production of reactive oxygen species induced by salt stress. Rice (Oryza sativa L.) genotypes BRS Bojuru and BRS Pampa, tolerant and sensitive to salinity, respectively, were subjected to 150 mM NaCl for 0, 6, 24, 48, and 72 h. A significant increase of superoxide anion and H₂O₂ and a decrease in malondialdehyde (MDA) content were observed in the tolerant genotype, whereas in the sensitive genotype, there was no change in superoxide anion content, reduced H₂O₂ content, and increased MDA content. The superoxide dismutase (SOD) activity increased significantly in both genotypes, and increases in amounts of transcript were observed for OsSOD3Cu/Zn and OsSODA1-Mn in the tolerant genotype and for OsSOD4-Cu/Zn, OsSOD3-Cu/Zn, OsSODCc1-Cu/Zn, OsSOD-Fe, and OsSODA1-Mn in the sensitive genotype. The activities of catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR) were not significantly and consistently changed, but OsCATA, OsAPX2 and OsGR1 were induced in both genotypes. OsCATB transcription was increased in the tolerant genotype and OsCATC and OsAPX3 in the sensitive genotype under salinity. It is concluded that OsAPX3, OsGR2, OsGR3, and OsSOD3-Cu/Zn genes are the most suitable to distinguish tolerant from sensitive genotypes under salt stress.

In chloroplasts and mitochondria, antioxidant mechanisms include the ascorbate-glutathione cycle, and monodehydroascorbate reductase (MDHAR) is important for regeneration of ascorbate (AsA) from monodehydroascorbate (MDHA). To improve detoxification of reactive oxygen species (ROS), we established a construct of the MDHAR gene from Brassica rapa fused to the targeting signal peptides of Pisum sativum glutathione reductase (GR), which was controlled by a stress-inducible SWPA2 promoter, and introduced this expression system into Arabidopsis thaliana. Transgenic (TG) plants overexpressing BrMDHAR targeted to chloroplasts and mitochondria through signal peptides showed an elevated MDHAR activity and an increased ratio of AsA to dehydroascorbate (DHA) when compared to wild-type (WT) plants under a freezing stress. These led to increased photosynthetic parameters, redox homeostasis, and biomass in TG plants when compared to the WT plants. Our results suggest that the overexpression of the BrMDHAR gene targeted to chloroplasts and mitochondria conferred an enhanced tolerance against the freezing stress, and a stress adaptation of dual-targeted BrMDHAR was better than that of single BrMDHAR.

Polyunsaturated fatty acids (PUFAs) affect diverse physiological processes and human health. Most cereals are poor in n-3 and n-6 PUFAs. Using biolistics, barley (Hordeum vulgare L. cv. Golden Promise) was transformed with an artificial gene encoding Δ⁶-desaturase (D6D) under an endosperm-specific promoter. This artificial gene was designed from the sequence of D6D of the filamentous fungus Thamnidium elegans, but codon usage was optimised for cereals. A signal sequence from the gene encoding for high molecular mass glutenin Dx5 was added to a destinate mature protein. Successful transformation was confirmed in T₀ plants at the genomic level and in T₁ seeds at the transcriptomic and metabolomic levels. Transformed plants produced up to 0.141 % of γ-linolenic acid (GLA) and 0.294 % of stearidonic acid (SDA) of the total amount of fatty acids in their grains. Although the content of these fatty acids was relatively low, the current study provides the first evidence that transgenic barley can be a source of GLA/SDA.

Transformation and high efficient regeneration of transgenic plants from the trimmed etiolated shoot/root region (TESRR) of Anliucheng sweet orange [Citrus sinensis (L.) Osb.] seedling was reported. A visual green fluorescent protein (GFP) marker gene was introduced to evaluate transformation efficiency by using the explants from TESRR and epicotyls. The transformation protocol was: infection 20 min, co-culture 3 d, selection culture 30 d, and rooting 15 d. Out of a total of 288 sprouted shoots obtained from TESRR, 34 shoots (11.8 %) yielded GFP expression. In contrast, only 2 (3.0 %) of the 67 sprouted shoots from epicotyl transformation yielded GFP expression. In all plants showing the green fluorescence an expected 500 bp GFP fragment was proved by PCR analysis. Southern blot analysis further confirmed the integration of GFP gene into citrus genome. Transgenic plantlets were obtained within 80 d using the TESRR, compared within 150 d by using epicotyls.

A gene encoding Mn-containing superoxide dismutase (Mn-SOD), designated as ThMSD, was cloned from salt cress (Eutrema halophilum) by reverse transcriptase - polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). The full length of ThMSD (acc. No. EF413171) is 1 047 bp with an open reading frame (ORF) of 693 bp. The deduced 231-amino acid polypeptide had a predicted molecular mass of 25.5 kDa, an estimated pI of 9.08, and a putative Mn-binding site. Recombinant ThMSD protein was expressed in Escherichia coli and characterized. The SOD activity of ThMSD was inactivated by sodium azide but not by potassium cyanide or hydrogen peroxide confirming that ThMSD is a Mn-SOD. Real-time PCR revealed that ThMSD was expressed in roots, rosette leaves, stems, stem leaves, flowers, and siliques. ThMSD mRNA reached the highest content in roots and its content increased when plants were treated with NaCl (in a concentration dependent manner), ABA, and subjected to drought. ThMSD was transformed into Arabidopsis and the stress tolerance properties of transgenic lines were assayed. The seeds of transgenic lines exhibited significantly higher germination rate under 100 and 150 mM NaCl than the wild type. The root growth of transgenic lines was affected less obviously than the wild type under 100 mM NaCl. The above results indicate that ThMSD played an important role in E. halophilum tolerance to environmental stresses, especially NaCl stress.

Agrobacterium-mediated genetic transformation is the most widely used technology to obtain overexpression of recombinant proteins in plants. Molecular events that occur within Agrobacterium during interactions with host plants have been studied extensively, and now we have a reasonable understanding the key factors involved in the regulation of T-DNA nuclear import and genomic integration. By contrast, very little is known about the events that take place in the host cells during genetic transformation by Agrobacterium. Here, we describe the plant-related factors including genotype, genes, proteins, competency of target tissues and phenolic compounds that participate in Agrobacterium-mediated genetic transformation and discuss their possible roles in this process. Because Agrobacterium probably adapts existing cellular processes for its life cycle, identifying the processes in host cells during Agrobacterium infection might contribute to better understanding of basic biological processes as cell communication, intracellular transport and DNA repair and recombination as well as to expanding the host range of Agrobacterium as a genetic engineering tool.

Plant glutathione peroxidases (GPX) catalyze the reduction of H₂O₂ or organic hydroperoxides to water, mitigating the toxicity of these compounds to cells. In rice plants, the GPX gene family is composed of five members that are distributed in a range of sub-cellular compartments including cytosol, mitochondria, chloroplasts, or endoplasmic reticulum. Of these, OsGPX1 and OsGPX4 are located in mitochondria and chloroplasts, respectively. To understand the role of these GPX in rice, the effect of knockdown of OsGPX1 and OsGPX4 in rice plants was evaluated. Our data show that OsGPX4 was essential for in vitro rice regeneration because no plants were obtained from calli carrying a hairpin construct against OsGPX4. Although the knockdown of OsGPX1 did not impair plant regeneration, the plants with silenced OsGPX1 (GPX1s plants) showed reduced shoot length and a reduced number of seeds compared to the non-transformed rice plants. These results indicate that OsGPX1 and OsGPX4 are essential for redox homeostasis which leads to normal growth and development of rice.

Abscisic acid (ABA) regulates plant responses to various environmental stresses. Oxidative cleavage of cis-epoxycarotenoids catalyzed by 9-cis-epoxycarotenoid dioxygenase (NCED) is the critical step in the biosynthesis of ABA in higher plants. Using a homologous cloning approach, a NCED-like gene (designated as TaNCED1) was isolated from wheat (Triticum aestivum). It contained an open reading frame of 1 848 bp and encodes a peptide of 615 amino acids. Multiple sequence alignments showed that TaNCED1 shared high identity with NCEDs from other plants. Phylogenetic analysis revealed that TaNCED1 was most closely related to a barley HvNCED1 gene. The predicted 3D structure of TaNCED1 showed high similarity with other homologues. Southern blot analysis indicated that TaNCED1 was a single copy in the genome of wheat. TaNCED1 was differentially expressed in various organs and the expression was up-regulated by low temperature, drought, NaCl, and ABA. Heterologous expression of TaNCED1 in tobacco (Nicotiana tabacum) significantly improved its drought tolerance. Under drought treatment, TaNCED1-overexpressing transgenic tobacco plants exhibited higher germination rate, higher relative water content, content of soluble sugars and of ABA when compared with the wild type plants.

Drought is a severe environmental constraint to plant productivity. Being a multidimensional stress, it triggers a wide variety of plant responses ranging from physiological, biochemical to molecular levels. One of the inevitable consequences of drought stress is an increase in reactive oxygen species (ROS) production in different cellular compartments, namely the chloroplasts and mitochondria. This enhanced ROS production is, however, kept under tight control by a versatile and cooperative antioxidant system that modulates intracellular ROS content and sets the redoxstatus of the cell. Furthermore, ROS production under stresses functions as an alarm signal that triggers defence or acclimation. Specific signal transduction pathways involve, e.g., H₂O₂ as a secondary messenger. ROS signalling under drought is linked to abscisic acid (ABA) and Ca²⁺ fluxes. At molecular levels, several drought-responsive genes, transcription factors, aquaporins, late embryogenesis abundant proteins, heat shock proteins, and dehydrins have been identified. This review discusses recent understanding on molecular responses and protective mechanisms of drought stress.

Osmotic stress causes a series of morphological, physiological, biochemical, and molecular changes that alters plant growth, development, and productivity around the globe. Phytohormones, nutrients, and transcription factors may induce adaptive responses to osmotic stress in plants. We evaluated the effect of osmotic stress induced by 18.5 % polyethylene glycol (PEG) or 100 mM NaCl on growth, content of abscisic acid (ABA), chlorophyll (Chl), sodium, and potassium, and the expression of multifunctional NAC transcription factors in rice cultivars (the salt-tolerant Cotaxtla and salt-sensitive Tres Ríos). The PEG and NaCl decreased shoot height and increased ABA content in both cultivars, and reduced root length in cv. Tres Ríos. The PEG increased Chl content in cv. Cotaxtla leaves. NaCl reduced shoot K⁺ content in cv. Tres Ríos and increased shoot and root Na⁺ content in both cultivars, thus resulting in a decreased K⁺/Na⁺ ratio. Of the 57 NAC genes evaluated, two of them were repressed (Os10g42130 and Os07g04560) and two other induced (Os02g34970 and OsNAC10) in cv. Cotaxtla in response to PEG, whereas three of them were repressed (Os10g42130, Os07g04560 and Os08g10080), and six induced (Os02g56600, Os02g34970, Os11g08210, Os05g34830, OsNAC6, and OsNAC10) in response to NaCl. In the cv. Tres Ríos, we found two genes repressed (Os10g42130 and Os07g04560), and five induced (Os08g33910, Os03g60080, Os06g51070, OsNAC6, and OsNAC10) in response to PEG, while only two genes were repressed (Os10g42130 and Os07g04560) but 13 induced (Os03g21060, Os08g39110, Os03g60080, Os01g15640, Os06g51070, Os09g33490, Os04g40130, Os12g29330, Os02g36880, Os11g08210, Os05g34830, OsNAC6, and OsNAC10) by NaCl. Osmotic stress affected more severely cv. Tres Ríos than cv. Cotaxtla plants. These different responses might be regulated by ABA and NAC transcription factors.

Brassinosteroids (Brs) have drawn wide attention due to their protective role against toxicity of heavy metals in plants. To better understand the role of Br in arsenic (As) and cadmium (Cd) uptake by rice plants, a hydroponic experiment was conducted to investigate the combined effect of 24-epibrassinolide (Br24) or 28-homobrassinolide (Br28) and iron plaque (IP) on As and Cd uptake and accumulation in rice seedlings. Six-week-old seedlings were sprayed with 0.2 or 0.02 μM Br24 or Br28 and grown in nutrient solution for 3 d, and then 20 or 60 mg Fe²⁺ dm^-3 (Fe20 and Fe60) was used to induce root IP formation for 3 d. These seedlings with or without Br and with or without IP were exposed to solution containing 0.5 mg dm^-3 As^III or Cd for 9 d. The results showed that rice growth decreased when Br24 were applied, but it increased when combination of Br24 and IP was applied. Fe concentrations in dithionite-citratebicarbonate (DCB) extracts were increased after 0.2 or 0.02 μM Br24 application in the absence of IP, but decreased by Br24 in the presence of IP. In the absence of IP, As and Cd content in leaves was significantly reduced by 0.02 μM Br24 and 0.2 μM Br28, respectively. The As content in leaves was also reduced by the combination of 0.02 and 0.2 μM Br28 and IP, and the Cd content in leaves was reduced by the combined effect of 0.2 μM Br24 and IP. These results indicate that Br24 and Br28 could impede As and Cd accumulation, and the interactions between Br and IP may have a potential in restricting the transport of As and Cd into rice shoots.

Sugars are important molecules that function not only as primary metabolites, but also as nutrients and signal molecules in plants. The sugar transport protein genes family SWEET has been recently identified. The availability of the Dendrobium officinale and Phalaenopsis equestris genome sequences offered the opportunity to study the SWEET gene family in this two orchid species. We identified 22 and 16 putative SWEET genes, respectively, in the genomes of D. officinale and P. equestris using comprehensive bioinformatics analysis. Based on phylogenetic comparisons with SWEET proteins from Arabidopsis and rice, the DoSWEET and PeSWEET proteins could be divided into four clades; among these, clade II specifically lacked PeSWEETs and clade IV specifically lacked DoSWEETs, and there were orthologs present between D. officinale and P. equestris. Protein sequence alignments suggest that there is a predicted serine phosphorylation site in each of the highly conserved MtN3/saliva domain regions. Gene expression analysis in four tissues showed that three PeSWEET genes were most highly expressed in the flower, leaf, stem, and root, suggesting that these genes might play important roles in growth and development in P. equestris. Analysis of gene expression in different floral organs showed that five PeSWEET genes were highly expressed in the column (gynostemium), implying their possible involvement in reproductive development in this species. The expression patterns of seven PeSWEETs in response to different abiotic stresses showed that three genes were upregulated significantly in response to high temperature and two genes were differently expressed at low temperature. The results of this study lay the foundation for further functional analysis of SWEET genes in orchids.

Heterotrimeric guanine-nucleotide-binding proteins (G-proteins) play important roles in signal transduction and regulate responses to various stresses. Although G-protein signaling pathways have been extensively identified and characterized in model plants, there is little knowledge in non-model and especially in woody plants. Mulberry is an economically and ecologically important perennial tree, which is adaptable to many environmental stresses. In this study, we identified and cloned six G-protein genes including one Gα, one Gβ, two Gγ, one RGS (regulator of G-protein signaling protein) and one RACK1 (receptor for activated C kinase 1) involved in G-protein signaling. Sequence and phylogenetic analysis revealed that Morus G-proteins are evolutionarily conserved compared with those of other plants. A real-time quantitative reverse transcription polymerase chain reaction analysis showed that Morus G-protein signaling genes were ubiquitously but differentially expressed in various tissues. The expression of all of these genes was affected by abiotic stresses and signal molecules, which indicated that Morus G-protein signaling may be involved in environmental stress and defense responses.

Elymus nutans is an important alpine perennial forage of the Pooideae subfamily, that can survive subzero temperatures. To understand the molecular mechanisms underlying cold tolerance in E. nutans, we performed the transcriptional analysis by RNA-Seq in two genotypes, the tolerant Damxung (DX) and the sensitive Gannan (GN), under cold stress. The new E. nutans transcriptomes comprised 200 520/200 836 and 181 331/211 973 transcripts in leaves/crowns of DX and GN, respectively. More cold-stress-related genes were identified in leaves than in crowns of both genotypes throughout the whole cold stress. The most prominent functional category in leaves of both genotypes at 3 h of stress was transcriptional regulation. Brassinosteroid and jasmonic acid mediated signalling pathways play central roles in regulating downstream protective responses in DX after 24 h of cold stress. Prolonged cold stress caused more severe transcriptome responses in crowns and leaves of DX compared to GN. The most significant transcriptomic changes in both genotypes were associated with the response to abiotic stresses and the oxidation-reduction processes, implying reprogramming of the cellular metabolism as an adaptation to cold stress. This study reveals mechanisms of genotype- and organ-specific cold stress response in E. nutans and thus provides a basis for future breeding strategies aimed at improving the tolerance of cold-sensitive plants.

Gibberellic acid (GA) promotes cotton fiber cell elongation under in vitro culture conditions and GA signal transduction is mediated by DELLA proteins. Our laboratory has cloned several DELLA genes from cotton and generated transgenic lines in Arabidopsis over-expressing the DELLA gene (GhRGL) and the DELLA-domain deletion mutant gene (Ghrgl). Comparative transcriptome profiling was performed on Arabidopsis transgenic lines over-expressing both GhRGL and Ghrgl. KEGG pathway analysis indicated that the differential genes participated in many important pathways, such as response to gibberellic acid, fatty acid metabolism, etc., and the expression patterns of 93 genes were validated by RT PCR in all transgenic lines and the wild-type plants. Gene ontology annotation identified that these genes were involved in sucrose synthesis, actin nucleation, root hair development, and various hormone signal responses.

WRKY transcription factors (TF) family is involved in a huge variety of plant processes, including seed germination, plant development, phytohormone signalling, and defence against both biotic or abiotic stresses. In this work, WRKY TF family has been characterized in citrus. In a first experiment, the relative expression of CsWRKYs was analyzed in shoots and roots of plants treated with abscisic acid (ABA), salicylic acid (SA), and methyl jasmonate (MeJA) under in vitro conditions. Expression of CsWRKYs was also determined in roots of commercial citrus rootstocks subjected to osmotic and salt stresses. A total amount of 50 CsWRKYs has been found and devided into different groups of WRKY family according to the WRKY domain sequences. In response to the applications of phytohormones, the highest differences were observed in roots, and it was found that ABA and SA generally repressed CsWRKYs expressions, but MeJA induced their overexpression. Osmotic stress repressed the expression of most of the CsWRKYs, whereas salt stress induced their expression. Moreover, salt stress induced higher increase in CsWRKY expressions in the tolerant rootstock Citrus macrophylla than in the sensitive rootstock Carrizo citrange, suggesting that these TFs may play an important role in response to this stress.

Sulfur dioxide is a widespread air pollutant and it also acts as a signaling molecule in various processes in mammals. However, the role of SO₂ in programmed cell death (PCD) in plants is unclear. Here we studied the role of SO₂ in gibberellin (GA)-treated wheat aleurone layers. The results showed that 100 μM SO₂ donor (NaHSO₃/Na₂SO₃) could effectively delay PCD and inhibit the coalescence of small protein storage vacuoles (PSVs) in aleurone cells treated with GA. Also, SO₂ could reduce the accumulation of hydrogen peroxide and superoxide anion in GA-treated aleurone layers. In this process, SO₂ could sustain higher activities of catalase, guaiacol peroxidase, ascorbate peroxidase, and superoxide dismutase and lower activities of lipoxygenase and polyphenol oxidase by comparing with GA alone. In addition, an induction of endogenous H₂S and NO was observed in SO₂-treated aleurone layers. The application of NO scavenger cPTIO could accelerate PCD in SO₂ or H₂S treated aleurone cells, suggesting that NO alleviated PCD by acting downstream of SO₂ and H₂S. In conclusion, these results imply that SO₂ could delay PCD in GA-treated wheat aleurone layers by enhancing cellular antioxidative capacity and H₂S/NO signals act downstream of SO₂.

Tomato (Solanum lycopersicum) plants have four fructokinase genes, SlFRK1-4. The SlFRK4 is expressed only in pollen whereas the other three are expressed in all plant parts. While SlFRK2 and SlFRK3 are involved in vascular tissue development and affects the shape, size, and cell-wall width of xylem vessels and xylem fibers, the role of SlFRK1 has not been studied previously. The current work investigates the expression of SlFRK1 using transgenic tomato plants expressing the β-glucuronidase reporter gene under the SlFRK1 promoter, as well as the role of SlFRK1 using transgenic plants with antisense suppression of SlFRK1. The SlFRK1 promoter is expressed primarily in vascular tissues and specific suppression of SlFRK1 reduces water transport in stems, but has no other anatomical or phenotypic effects. Combined suppression of SlFRK1 and SlFRK2 severely inhibited plant growth and an anatomical analysis revealed a reduction in secondary xylem area and distorted phloem fibers characterized by thin cell walls and reduced lignification. The results suggest that SlFRK1 is involved in vascular tissue development and hydraulic conductivity in tomato plants and that SlFRK1 is important for normal phloem fiber development, together with SlFRK2.

DNA methylation is one of the epigenetic mechanisms regulating gene expression in plants in response to environmental conditions. In this study, analysis of methylation patterns was carried out in order to assess the effect of water stress in two contrasting wheat genotypes using methylation-sensitive amplified polymorphism (MSAP). The results revealed that demethylation was higher in drought-tolerant genotype (C306) as compared to drought-sensitive genotype (HUW468) after experiencing drought stress. Comparisons of different MSAP patterns showed a high percentage of polymorphic bands between tolerant and susceptible wheat genotypes (from 74.79 % at anthesis to 88.89 % at tillering). Furthermore, differential DNA methylation in roots and leaves also revealed tissue-specific methylation of genomic DNA. Interestingly, 54 developmental stage-specific bands and 23 bands that were found contrasting between these two wheat genotypes were detected. Furthermore, a few sites with stable DNA methylation differences were identified between drought-tolerant and drought-sensitive cultivars, thus providing genotype-specific epigenetic markers. These results not only provide data on differences in DNA methylation changes but also contribute to dissection of molecular mechanisms of drought response and tolerance in wheat.

Uridine diphosphate glucose dehydrogenase (UGDH) plays an important role in biosynthesis of hemicellulose by catalyzing oxidation of UDP-glucose (UDP-Glc) to UDP-glucuronate (UDP-GlcA), a key sugar nucleotide involved in biosynthesis of the plant cell wall. In this study, a UGDH ortholog referred to as LgUGDH was isolated from Larix gmelinii using PCR and rapid amplification of cDNA ends techniques. Real-time PCR shows that the LgUGDH gene was expressed primarily in larch stems in addition to its roots and leaves, and Southern blot analysis indicates that UGDH is encoded by two paralogous genes in L. gmelinii. Overexpression of LgUGDH increased the content of soluble sugars and hemicelluloses and enhanced vegetative growth and cold tolerance in transgenic Arabidopsis thaliana. These results reveal that L. gmelinii UGDH participates in sucrose/polysaccharide metabolism and cell wall biosynthesis and may be a good candidate gene for enhancing plant growth, cold tolerance, and hemicellulose content.

We studied changes in physiological parameters of whole leaves and in antioxidant protection of chloroplasts during ageing and senescence of tobacco (Nicotiana tabacum L. cv. Samsun NN) leaves with enhanced cytokinin oxidase/dehydrogenase activity (CKX) or without it (WT). Old leaves of CKX plants maintained higher pigment content and photosystem 2 activity compared to WT leaves of the same age. Chloroplasts of old CKX plants showed better antioxidant capacity represented by higher superoxide dismutase, dehydroascorbate reductase and glutathione reductase activities.

In Arabidopsis, EXPORTIN1A (HIT2/XPO1A) and EXPORTIN1B (XPO1B) mediate the translocation of nuclear export sequence (NES)-bearing proteins from nucleus to cytoplasm. However, a mutation in HIT2/XPO1A but not in XPO1B induces sensitivity to high irradiance (HI). Arabidopsis thaliana heat stress elements A4a and A5 (AtHsfA4a and AtHsfA5) are involved in plant responses to HI and possess NESs; therefore, their nucleo-cytoplasmic partitioning was analyzed. In wild-type and xpo1b mutant cells, AtHsfA4a normally remained in the cytoplasm but became concentrated in the nucleus following exposure to HI, whereas AtHsfA5 was constitutively distributed in both cytoplasm and nucleus. However, in hit2/xpo1a mutant, AtHsfA4a and AtHsfA5 were always confined to the nucleus, regardless of the irradiance. Although AtHsfA4a can enhance the ability of plants to scavenge H₂O₂, and AtHsfA5 is a repressor of AtHsfA4a, athsfa5 but not athsfa4a mutant plants exhibited HI sensitivity. Additionally, athsfa4a plants expressing AtHsfA4aΔNES were sensitive to HI, but athsfa5 plants expressing AtHsfA5ΔNES were not. Meanwhile, hit2/athsfa4a double mutant was more tolerant to HI than hit2. These results indicate that both AtHsfA4a and AtHsfA5 were HIT2/XPO1A-specific substrates. Long-term accumulation of AtHsfA4a contributed to the hit2 HI-sensitive phenotype independent of the scavenging ability of H₂O₂, and the presence of AtHsfA5 could mitigate this adverse effect.

Phosphites (Phis), inorganic salts of phosphorous acid, have shown to be effective in protection of plants against biotic stress. Recently, we have described that potassium phosphite (KPhi) induces tolerance to UV-B radiation in potato. To counteract the harmful effect of UV radiation, plants accumulate UV-screening compounds, such as flavonoids, sinapate ester, and lignin. In previous work, we have shown an increase in guaiacol peroxidase (POD) activity in plants pretreated with KPhi and further exposed to UV-B radiation. In order to continue with this study, the expression of different enzymes and components involved in cell wall reinforcement were analyzed. An isoform of POD induced by KPhi was analyzed by isoelectric focusing and further identified as suberization-associated anionic peroxidase (POPA) by mass spectrometry. In addition, other enzymes participating in lignin biosynthesis, like caffeoyl-CoA O-methyltransferase (CCoAOMT), determined by accumulation of transcripts, and laccase activity, visualized in zymogrames, were increased by KPhi treatment previous to UV-B exposure. Further, the accumulations of extensin (EXT) transcripts and of conjugated polyamines (PAs) were increased by KPhi treatment previous to UV-B exposure. All these results suggest cell wall reinforcement in leaves due to KPhi pretreatment followed by UV-B exposure.

Root hairs play important roles in plant nutrient and water acquisition. To better understand the genetic mechanism controlling root hair development in rice (Oryza sativa L.), a rice mutant with root hair defects was isolated and characterized. Cryo-scanning electron microscope (SEM) showed that the density and length of root hairs in the mutant were significantly reduced compared to wild type (WT). Map-based cloning and complementation test revealed that the mutation occurred in a NADPH oxidase gene OsNOX3 (LOC_Os01g61880). The OsNOX3 displays high sequence similarity with the previously characterized NOX genes RTH5 in maize and RHD2 in Arabidopsis, which play critical roles in root hair development. Expression pattern analysis indicated that OsNOX3 is expressed in various tissues throughout the plant with high expression in roots and root hairs. Subcellular localization analysis confirmed that OsNOX3 is located on the plasma membrane. Staining assays showed that the content of superoxide and hydrogen peroxide are significantly reduced in root hair tips of Osnox3 when compared to WT. Our results showed critical roles of OsNOX3 in regulating both root hair initiation and elongation in rice, which is similar to RTH5 but different from RHD2, confirming the difference of genetic mechanisms regulating root hair morphogenesis in monocot and dicot plants.

Gibberellins (GAs) are a large family of tetracyclic diterpenoids, controlling important aspects of growth and development throughout the plant life cycle. To explore the possibility that gibberellin A₃ (GA₃) signalling induces epigenetic alteration(s), we carried out a field experiment study using Nicotiana tabacum as a model system. The GA₃ application on leaves resulted in increased plant-height, foliage density, leaf cell area, and trichome density. The plants exposed to GA₃ also exhibited: 1) increased chromatin de-condensation, 2) reduced global DNA methylation, 3) reduced DNA methyltransferases (NtDNMTs) activities accompanied by decreased amounts of NtMET1 and NtCMT3 transcripts, and 4) partial restoration of phenotype and expression of epigenetically silenced reporter transgene. Based on these observations, we propose that GA₃ application induces complex epigenetic re-programming, which may lead to distinct developmental phenotypes. These results could provide an important insight for future studies on epigenetic mechanism(s) in other important crops.