Uridine diphosphate glucose dehydrogenase (UGDH) is a key enzyme in the hemicellulose and pectin biosynthesis pathway and participates in the regulation of growth and development in plants. In this study, we isolated a BpUGDH gene from paper mulberry (Broussonetia kazinoki × Broussonetia papyifera) and analyzed its function and expression characteristics. The results show that the BpUGDH was expressed in all organs of paper mulberry with a higher expression in stems than in leaves and roots. A pBpUGDH::GUS gene construct was highly expressed in transgenic Arabidopsis thaliana seedlings, and its expression was induced by a low temperature, methyl jasmonate, gibberellin A₃, ethylene, and auxin. The overexpression of BpUGDH increased the soluble sugar content, promoted the accumulation of hemicellulose, and enhanced the vegetative growth of transgenic plants. These results provide a basis for regulating the growth and adaptability of paper mulberry and improving its utilization value via genetic modification of the BpUGDH gene.

Wheat (Triticum aestivum L.) is leading cereal crop worldwide, but its yield is highly affected due to various diseases, especially Fusarium head blight (FHB), which affects the metabolism of plants. The present study was conducted at the Agricultural Institute Osijek using three winter wheat cultivars (Apache, Bezostaya1, and U1) during 2016/2017. The objectives of our studies were to examine differences in physiological characteristics of FHB resistance among wheat cultivars in the early stage of infection. The FHB incidence and severity was the highest in 'Bezostaya1'. Results suggest that activation of some anti-oxidative enzymes in the first 2 h after Fusarium attack was not efficient to prevent disease. 'Apache', which revealed an average FHB incidence, efficiently activated defence response through phenol metabolism elevation. The most effective defence response trough activation of anti-oxidative enzymes triggered by H₂O₂ was revealed in 'U1', which resulted in a minimal FHB incidence and disease severity. The obtained results confirm differences in defence strategies of wheat genotypes.

In plants, auxin/indoleacetic acid (Aux/IAA) proteins are transcriptional regulators, which regulate developmental process and responses to phytohormones and stress treatments. A previous study has shown that the rice Aux/IAA transcription factor gene OsIAA18 is induced by salt and osmotic stresses. However, little is known about the regulatory functions of this gene. In this study, the OsIAA18 gene was successfully cloned from rice. Subcellular localization analysis in onion epidermal cells indicated that OsIAA18 was localized to the nucleus. Expression analysis in yeast showed that the full length OsIAA18 exhibited transcriptional activation. Heterologous expression of OsIAA18 significantly enhanced salt and osmotic tolerance in transgenic Arabidopsis plants. Real-time quantitative PCR analysis showed that constitutive expression of OsIAA18 up-regulated genes involved in abscisic acid (ABA) biosynthesis, proline biosynthesis, stress responses, and reactive oxygen species scavenging under salt and osmotic stresses. Enzymatic analyses found that the transgenic plants had higher 9-cis-epoxycarotenoid dioxygenase, pyrroline-5-carboxylate synthase, superoxide dismutase, and peroxidase activities than wild-type plants under salt and osmotic stresses. In the transgenic plants, ABA and proline content significantly increased, whereas H₂O₂ and malondialdehyde content significantly decreased. In addition, the transgenic plants had also a lower electrolyte leakage and water loss rate. These overall results indicate that the OsIAA18 gene is involved in enhancing salt and osmotic tolerance in transgenic Arabidopsis plants. The OsIAA18 gene has a potential to be used to enhance the tolerance to abiotic stresses in other plant species.

Festulolium cultivars are widely utilized in Lithuania because they are persistent under abiotic stresses and are high yielding. However, changing climate challenges the existing Festulolium cultivars to adapt to new growing conditions and still maintain the yield. In this study, we aimed at evaluating the yield stability of two Festulolium cultivars in field trials under fluctuating Lithuanian conditions. The mean total dry matter yield (DMY) of both Festulolium cultivars fluctuated greatly between the years and ANOVA analysis showed a significant effect of environment on total DMY as well as DMY of each cut, but the genotype × environment interaction was not significant. There was a high difference between the total DMY of 1^st year and 2^nd year of use of plots in each year of observation. The highest DMYs were harvested in the years 2015 and 2016. Dry matter yield of the 1^st cut was the largest component of the total DMY for most of the years. The plants overwintered the first winter after sowing very well over the whole study period, resulting in excellent spring growth. The winter survival scores of 2^nd year of use of plots were lower than 1^st year of use and strongly correlated with the 1^st cut DMY of 2^nd year of use (r = 0.81). Spring growth of plants at 2^nd year of use was poorer, the correlation between winter survival and spring growth of 2^nd year of use was 0.62. The scores of regrowth after the cuts of 1^st and 2^nd years of use were very similar for most of the experimental years and moderately correlated with the sum of DMYs after cuts (r = 0.55 and r = 0.5, respectively).

Pyramiding transgenes of interest is one of the strategies to engineer multiple stress resistance in crop plants. Transgenic plants which stably express different genes can be hybridized to bring these genes together in one plant. Transgenic rice (Oryza sativa L. cv. ASD 16) plants harbouring genes Xa21 (conferring bacterial blight resistance), tlp (conferring resistance to sheath blight), or gna (conferring resistance to brown planthopper) were used in hybridization experiments. Sexual hybridization was carried out in two different gene combinations: Xa21 × gna and tlp × gna. Molecular analyses were carried out to confirm the presence of transgenes. In F₁ generation, lines harbouring either gene in each of the cross-combination were selected and forwarded to F₂ generation. The presence of genes in F₂ generation was confirmed by PCR, Southern blot hybridization, and Western blotting. The F₂ progeneis harbouring Xa21 and gna exhibited resistance against bacterial blight and moderate resistance against brown planthopper. Similarly, the F₂ lines of tlp and gna combination provided resistance against sheath blight and moderate resistance against brown planthopper. The level of resistance observed in pyramided lines for insect or pathogens was comparable to the resistance observed in their parental lines. Our study shows that pyramiding genes by hybridization between transgenic plants could be one of the strategies to develop cultivars with multiple biotic stress resistances.

Mannose (MAN), an important monosaccharide, contributes to coping with abiotic stresses in plants. Objectives of this study were to examine whether exogenous MAN (30 mM) could significantly increase drought tolerance and further to reveal MAN-regulated tolerance mechanism in white clover under osmotic stress induced by 18 % (m/v) polyethylene glycol 6000 for 10 d in controlled growth chambers. Results show that the application of MAN significanlty alleviated stress damage and the inhibition of growth and photosynthesis in white clover under osmotic stress. The MAN-induced increase in endogenous MAN content and the accumulation of organic osmolytes (proline and water soluble sugars) could be responsible for a lower osmotic potential (OP) in white clover. The exogenous application of MAN also enhanced antioxidant enzyme (superoxide dismutase, peroxidase, ascorbate peroxidase, dehydroascorbate reductase, and glutathione reductase) activities and maintained ascorbic acid content in white clover during osmotic stress. As concern chlorophyll (Chl) metabolism, the MAN-treated plants showed significantly higher transcription of genes involved in Chl synthesis Mg-chelatase and protochlorophyllide reductase and lower transcription of pheophorbide a oxygenase and chlorophyllase related to Chl degradation and also a senescence associated gene 101 than untreated plants. In addition, the MAN application increased transcription of SK2-, Y2K-, and Y2SK-type dehydrin genes, and dehydrin b in leaves of white clover under osmotic stress. These results indicate that MAN plays important roles in drought tolerance not only acting as a compatible solute for OP but also delaying leaf senescence through enhancing antioxidant metabolism, decreasing Chl degradation, and increasing transcription of dehydrin genes contributing to enhanced drought tolerance in white clover.

Interspecific and intergeneric hybridization within the Festuca-Lolium complex is frequently used in forage plant breeding. However, little is known about the natural occurrence and competitiveness of such hybrids. We collected naturally formed hybrids between Festuca apennina, Festuca pratensis, and Lolium perenne in different habitats of Switzerland and the British Isles and studied their origin, the ease of their spontaneous formation, and their competitiveness with parental species. A special attention was paid to the largely sterile triploid forms and their rare sexual progeny. The triploid hybrid F. apennina × F. pratensis proved to be widespread and often highly competitive in Swiss permanent pastures. The majority of these hybrids originated from F. apennina as the seed parent although little or no F. apennina grew nearby. In an experimental setting with ample F. pratensis pollen provided by neighbouring plants, up to 20 % of seeds from open pollinated F. apennina plants were interspecific hybrids; among seeds collected in natural habitats, only 0.35 % were hybrids. At an experimental site at 1 000 m altitude, these triploid hybrids grew much more vigorously than corresponding tetraploid pure F. apennina, confirming their great competitiveness at such altitudes in permanent grasslands. The triploid hybrids were only marginally fertile suggesting that vegetative propagation by rhizomes is the cause of their competitive success in grassland. Moreover, triploid progeny retained the chromosome constitution of their mother plants indicating the possibility of apomixis. Natural triploid F. pratensis × L. perenne hybrids were partially female fertile (a seed set of 0.1 % or less) whereas diploid hybrids did not produce any viable seeds. Progenies of these triploids showed considerable chromosome alterations, such as loss of a genome or recombination due to homoeologous pairing, and only rarely the chromosome constitution of the triploid mother plant was retained. It was concluded that natural triploid interspecific hybrids could expand the range of their progenitor species and might function as bridges transferring genes between them.

Plants face variable environmental stresses that negatively affect plant growth and productivity. The multiplicity of responses is an important aspect of the complexity of stress signalling. Abscisic acid (ABA) is a broad-spectrum phytohormone involved not only in regulating stomatal opening, growth and development but also in coordinating various stress signal transduction pathways in plants during abiotic stresses. The both ABA-dependent and ABA-independent signal transduction pathways from stress signal perception to gene expression involve different transcription factors such as DREB, MYC/MYB, AREB/ABF, NAM, ATAF1,2, CUC and their corresponding cis-acting elements DRE, MYCRS/MYBRS, ABRE, NACRS. Genetic analysis of ABA mutants has given insight that ABA-dependent and ABA-independent pathways for osmotic stress and cold stress interact and converge. This review focuses on ABA-dependent and ABA-independent transcriptional components and cascades, their specificity and crosstalk in stress gene regulation.

Tospoviruses are devastating plant viruses causing severe economic losses in a diverse range of crops worldwide. Here, we describe the development and evaluation of an RNA interference (RNAi) broad-spectrum virus resistance strategy based on a unique and short hairpin-RNA-generating construct (pNhpRNA). This construct was designed from a region of the nucleocapsid gene (N) of Tomato spotted wilt virus (TSWV) that showed a high sequence identity to the corresponding region in the related species Groundnut ringspot virus (GRSV) and Tomato chlorotic spot virus (TCSV). To test the effectiveness of the pNhpRNA construct, we developed a silencing reporter assay based on three fusion proteins in which the complete viral N gene sequence from each of the three tospoviruses was fused in frame to the green fluorescent protein (GFP) sequence. Co-agroinoculation of these constructs with pNhpRNA into leaves of Nicotiana benthamiana resulted in a strong silencing phenotype determined by GFP decay and suppression of the three N genes at the RNA and protein levels. To test the potential of the pNhpRNA construct to generate virus-resistant plants, we infiltrated the whole shoots of N. benthamiana with pNhpRNA. When these infiltrated plants were mechanically inoculated with the mentioned viruses 100, 70, and 60 % resistance phenotypes to TSWV, GRSV, and TCSV, respectively, were observed. The induction of a broad tospovirus resistance with a simple construct and a minimized off-target effect are the main contributions of pNhpRNA.

The responses of antioxidant enzymes (AOE) ascorbate peroxidase (APX), glutathione reductase (GR), superoxide dismutase (SOD), and catalase (CAT) in soluble protein extracts from leaves and roots of tobacco (Nicotiana tabacum L. cv. Samsun NN) plants to the drought stress, salinity and enhanced zinc concentration were investigated. The studied tobacco included wild-type (WT) and transgenic plants (AtCKX2) harbouring the cytokinin oxidase/dehydrogenase gene under control of 35S promoter from Arabidopsis thaliana (AtCKX2). The transgenic plants exhibited highly enhanced CKX activity and decreased contents of cytokinins and abscisic acid in both leaves and roots, altered phenotype, retarded growth, and postponed senescence onset. Under control conditions, the AtCKX2 plants exhibited noticeably higher activity of GR in leaves and APX and SOD in roots. CAT activity in leaves always decreased upon stresses in WT while increased in AtCKX2 plants. On the contrary, the SOD activity was enhanced in WT but declined in AtCKX2 leaves. In roots, the APX activity prevailingly increased in WT while mainly decreased in AtCKX2 in response to the stresses. Both WT and AtCKX2 leaves as well as roots exhibited elevated abscisic acid content and increased CKX activity under all stresses while endogenous CKs and IAA contents were not much affected by stress treatments in either WT or transgenic plants.

A new member of the APETALA2/ethylene responsive element binding protein (AP2/EREBP) transcription factor family, HhDREB2, was isolated from Halimodendron halodendron. Based on the similarity of the AP2/ERF domain, HhDREB2 was classified into A-5 group of the DREB subfamily. The expression of HhDREB2 gene was induced by drought, high salinity, and low temperature, but not by exogenous plant hormones. Trans-activity assay demonstrated that HhDREB2 gene encodes a transcription activator. Furthermore, over-expression of HhDREB2 gene under the stress-inducible rd29A promotor in Arabidopsis resulted in enhanced tolerance to salt and drought stresses. The overall results reveal that HhDREB2 functioned as important transcription factor in regulation of stress-responsive signaling in plants and may be used for improving plant tolerance to abiotic stresses.

Germin and germin-like proteins (GLPs) are a broad and diverse family of developmentally regulated proteins widely distributed in plants. Oryza sativa L. harbours a large family of GLPs and serves as a good model for their study. In the present study, a germin-like protein gene (OsRGLP1) of rice origin was characterized by its heterologous expression in tobacco. The real-time PCR established almost a uniform expression of OsRGLP1 in leaves, stem, and roots of T₁ Nicotiana tabacum cv. Samsun. Although no morphological difference was apparent between T₀ transgenic and wild-type plants, leaves of mature transgenic plants showed necrotic lesions associated with an elevated content of H₂O₂, which was evidenced by in situ 3,3'-diaminobenzidine staining. A significantly higher activity of heat resistant superoxide dismutase (SOD) was observed in the transgenic plants as compared to the wild-type. The SOD activity in the transgenic plants was insensitive to potassium cyanide and sensitive to H₂O₂.

MAT (multi-auto-transformation) vector system has been one of the strategies to excise the selection marker gene from transgenic plants. Agrobacterium tumefaciens strain EHA105 harboring an ipt-type MAT vector, pNPI132, was used to produce morphologically normal transgenic Petunia hybrida 'Dainty Lady' employing isopentenyl transferase (ipt) gene as the selection marker gene. β-glucuronidase (GUS) gene was used as model gene of interest. Infected explants were cultured on Murashige and Skoog (MS) medium without plant growth regulators (PGR) and antibiotics. Shoots showing extreme shooty phenotype (ESP) were produced from the adventitious shoots separated from the explants. Visual selection was carried out until production of morphologically normal shoots (approximately 4 months after infection). Histochemical GUS assay detected GUS gene in both ESP and normal shoots. PCR analysis confirmed the presence of model gene (GUS gene) and excision of the selection marker (ipt) gene in the normal transgenic plants. The insertion sites (1-3 for ipt gene and 1-2 for GUS gene) were detected by Southern blot analysis using DIG-labeled probes of both genes. These results show that ipt-type MAT vector can be used successfully to produce marker-free transgenic Petunia hybrida plants on PGR- and antibiotic-free MS medium.

The halophyte Thellungiella salsuginea is a new model plants due to its small genome size, short life cycle, and copious seed production. Although T. salsuginea shares a high sequence identity with its close relative Arabidopsis thaliana, it shows a greater tolerance to salinity, drought, freezing, heat, and cold. To elucidate the mechanism of abiotic stress resistance in T. salsuginea, we characterized its cytosolic Apx1 gene (TsApx1) and established A. thaliana transgenic lines overexpressing TsApx1. Under 300 mM NaCl, the content of H₂O₂, malondialdehyde, and proline were lower and the activities of superoxide dismutase, catalase, glutathione peroxidase, and ascorbate peroxidase were all higher in the transgenic plants overexpressing TsApx1 (35S:TsApx1-GFP) than in the wild-type plants. The atapx1 mutant plants of A. thaliana had a NaCl/mannitol-sensitive phenotype. The ectopic expression of TsApx1 in the atapx1 mutant effectively remedied the phenotype. These results suggest that TsApx1 plays an important role in scavenging reactive oxygen species in the cytoplasm under salinity or drought. Although TsApx1 in T. salsuginea was constantly expressed at a high level, this gene was clearly inducible. In summary, the high constitutive expression and rapid induction of TsApx1 may contribute to the tolerance to abiotic stresses in T. salsuginea.

In plant cells, anion channels and transporters are essential for key functions. Members of the chloride channel (CLC) family located in intracellular organelles are required for anion accumulation, pH adjustment, and salt tolerance. Here, we cloned a maize (Zea mays L.) CLC gene, named ZmCLC-d, and found that its transcription was up-regulated under cold, drought, salt, and heat stresses, and after hydrogen peroxide (H₂O₂) and abscisic acid (ABA) treatments. The overexpression of ZmCLC-d in Arabidopsis conferred tolerance to cold, drought, and salt stresses; this tolerance was primarily displayed by an increased germination rate, root length, plant survival rate, antioxidant enzyme (catalase, peroxidase, and superoxide dismutase) activities, and a reduced accumulation of Cl^- in transgenic plants as compared with wild type (WT) plants. The accumulation of H₂O₂ and superoxide anion in leaves of the ZmCLC-d-overexpressing plants is much less than that of the WT plants. The expressions of some stress related genes, such as CBF1, CBF2, CBF3, DREB2A, and RCI2A, increased to a greater extent in the ZmCLC-d-overexpressing plants than in the WT. Our results strongly suggest that ZmCLC-d played an important role in stress tolerance.

Previously, the ascorbate peroxidase (APX1) activity and gene expression in Chinese cabbage (Brassica campestris, Bc) heat-tolerant cv. ASVEG2 were found to be significantly higher than in heat-sensitive cv. RN720 under a heat stress. Furthermore, BcAPX2 and BcAPX3, isoforms of BcAPX1, were cloned in this study. Our objective was to transfer BcAPX cDNA under the control of the ubiquitin promoter to Arabidopsis via Agrobacterium tumefaciens strain GV3101. We found that BcAPX genes were overexpressed in transgenic Arabidopsis, and the expression of APX, and the APX activity in transgenic lines were higher than in non-transgenic (NT) plants under high temperatures. The chlorophyll content and the germination rate were significantly higher, and the malondialdehyde content was lower in BcAPX1-3, 2-1, and 3-5 lines subjected to the heat-stress treatment than those in the NT plants. Compared to the NT plants, a lower heat-induced H₂O₂ accumulation was detected by diaminobenzidine staining in leaves of the transgenic lines with a high APX activity indicating that the overexpression of BcAPX in Arabidopsis could enhance heat tolerance by eliminating H₂O₂.

Saussurea involucrata Kar. et Kir. tolerates severe abiotic stresses including cold, and the level of membrane fatty acid desaturation is associated with its cold acclimation. We discovered and characterized a full-length cDNA of stearoyl acyl-carrier-protein desaturase (sikSACPD) which encodes a protein consisted of 396 amino acids. A sequence alignment of the SikSACPD protein showed that it shares 91 and 86 % identity with the SACPDs of Carthamus tinctorius and Helianthus annuus, respectively. Semi-quantitative RT-PCR showed that the expression of sikSACPD increased in S. involucrata leaves as the temperature decreased from 20 to -10 °C. Agrobacterium tumefaciens was used to transform fatty acid biosynthesis 2 (FAB2):SikSACPD and FAB2:FAB2 constructs into tobacco to investigate resistance to a freezing stress and fatty acid composition of the transgenic plants. The FAB2:SikSACPD transgenic plants showed a slightly more resistance to the freezing stress than the FAB2:FAB2 transgenic plants and the wild-type. The proportion of oleic acid (C18:1) in the leaves of SikSACPD transgenic tobacco increased from approximately 5 to 20 % compared with the leaves of non-transgenic tobacco when both were exposed to cold stress treatments. This study demonstrates that the SikSACPD transgene, when expressed in tobacco, conferred a higher cold tolerance in comparison with that observed in non-transgenic tobacco. Thus, this gene may be a candidate for enhancing cold tolerance in other crop plants.

Iron deficiency chlorosis occurs frequently in calcareous soils. The transformation of plants with ferric chelate reductase genes (FROs) provides a potential strategy to alleviate plant chlorosis under iron deficiency. A CjFRO2 gene isolated from Citrus junos Sieb. ex Tanaka was introduced into Poncirus trifoliata (L.) Raf via Agrobacterium-mediated transformation. The transgene integration and expression were confirmed by PCR, Southern blot, and real-time PCR analyses. Hydroponic- and soil-grown transgenic plants were tested for their tolerance to iron deficiency. Compared with nontransgenic (NT) P. trifoliata plants, a rhizosphere acidification capacity in the transgenic lines increased, and a ferric chelate reductase activity in roots was up to 3.39- and 2.93-fold higher in a hydroponic solution and soil, respectively. A transgenic line TO-8, which reacted similarly in hydroponics and soil, appeared tolerant to the iron deficiency. Its leaf chlorophyll and ferrous ion content was significantly higher than in NT. These results indicate that tolerance to the iron deficiency in P. trifoliata could be improved through the genetic engineering.

Oxidative stress plays an important role in plant ageing and in response to different stresses. Oxidative DNA damage, unless repaired, may have detrimental consequences and increase genetic instability. Therefore, we determined the role of heat-shock induced oxidative stress on induction and repair of DNA damage in relation to oxidative stress tolerance in senescent tobacco plants. One-month-old (young) and three-month-old (senescent) plants were exposed to 42 °C for 2 and 4 h and left to recover at 26 °C for 24 and 72 h. The progression of senescence was characterized by the lower soluble protein and malondialdehyde content compared to young plants. Immediately after the heat shock, an increase in lipid peroxidation and guaiacol peroxidase activity, as well as DNA damage measured by the Comet assay were induced to higher extent in the young plants than in the senescent ones compared to their respective controls. Moreover, after 24-h recovery, the DNA damage further increased in the young plants whereas tendency of DNA repair was observed in the senescent plants. Upon 72-h recovery, no significant differences were noticed in all parameters studied (regardless of plant age) compared to the controls. The random amplified polymorphic DNA (RAPD) analysis confirmed genetic stability of the tobacco plants during the heat-shock exposures as well as the subsequent recovery periods.

To understand the protective roles of salicylic acid (SA) under high light, we investigated oxidative damage of Arabidopsis thaliana under high light in the presence or absence of SA. The results indicate that the high light led to an increase in the levels of proline, soluble sugars, reactive oxygen species, malondialdehyde, and electrolyte leakage, and a decrease in stomatal conductance (gs). Activities of six antioxidant enzymes increased significantly under the high light for 1 h. However, the high light for 3 h decreased the activities of peroxidase, superoxide dismutase, and catalase. In addition, we found that exogenous SA effectively improved antioxidant enzyme activities and significantly alleviated ROS accumulation and cell death in A. thaliana under the high light. Therefore, our results show that the high light caused a severe oxidative damage, and SA effectively alleviated the adverse effects of the high light on the plants by regulating the antioxidative defense system.

In non-heading Chinese cabbage, the yield relies mostly on the health of leaves, which can be heavily impacted by turnip mosaic virus (TuMV). The virions or viral ribonucleoprotein complexes are transported through the phloem and xylem. Plasmodesmata are indispensable because they traverse cell walls and connect companion cells, allowing virus particles long distance movement. However, which complexes and genes participate in this process is still unknown. Plants can activate defense mechanisms and apply disease resistance genes to respond to pathogen attacks. In this study, we collected the stems and petioles infected by TuMV for 7 d (TuMV-7), 14 d (TuMV-14), and 21 d (TuMV-21). Using isobaric tags for relative and absolute quantification-based proteomic technology, 6 043 distinct proteins were identified and 323, 240, 285, 203, 253, and 363 differentially expressed proteins were found in the comparable pairs of TuMV-7/control, TuMV-14/TuMV-7, TuMV-14/control, TuMV-21/TuMV-7, TuMV-21/TuMV-14, and TuMV-21/control, respectively. We performed a functional annotation analysis of all identified proteins and a functional enrichment analysis of all differentially expressed proteins. The results indicated that the long distance movement of TuMV involved many complex regulatory pathways. The respective proteins were related to those occurring in plasmodesmata and to Ca²⁺ transporters. Further, we also found proteins related to heat shock proteins, pathogenesis-related proteins, and proteins scavenging reactive oxygen species.

Salinity adversely affects plants resulting in disruption to plant growth and physiology. Previously, it has been shown that these negative effects can be alleviated by various exogenous polyamines. However, the role of spermidine (Spd) in conferring salinity tolerance in sorghum is not well documented. The effect of exogenous Spd on the responses of sweet sorghum (Sorghum bicolor L.) seedlings to salt stress (150 mM NaCl) was investigated by measuring photosynthetic carbon assimilation, Calvin cycle enzyme activities, and the the expression of respective genes. Application of 0.25 mM Spd alleviated the negative effects of salt stress on efficiency of photosystem II and CO₂ assimilation and increased the activities of ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) and aldolase. Salt stress significantly lowered the transcriptions of genes encoding Rubisco large subunit, Rubisco small subunit, 3-phosphoglyceric acid kinase, glyceraldehyde-3-phosphate dehydrogenase, triose-3-phosphate isomerase, fructose-1,6-bisphosphate aldolase, fructose-1,6-bisphosphate phosphatase, and sedoheptulose-1,7-bisphosphatase. However, transcriptions of genes encoding phosphoribokinase and Rubisco were up-regulated. The Spd application enhanced expressions of most of these genes. It appears Spd conferred salinity tolerance to sweet sorghum seedlings by enhancing photosynthetic efficiency through regulation of gene expressions and activities of key CO₂ assimilation enzymes.

Cannabis sativa, an annual herbaceous plant, produce wide variety of secondary metabolites among which delta-9-tetrahydrocannabinol (THC) is the most important one. The dissection of biosynthetic pathway(s) of this compound and its regulation by transcription factors (TFs) is an important prerequisite for efficient biotechnological manipulation of its secondary metabolome. A polyketide synthase (PKS) of C. sativa catalyzes the first step of cannabinoid biosynthesis, leading to the biosynthesis of olivetolic acid. Cloning and analysis of PKS promoter based on online PLACE, PlantCARE, and Genomatix Matinspector professional databases, indicated that PKS promoter consisted of cis-elements such as TATA-box, CAAT-box, W-box, Myb-box, E-box, and P-box. Plant expression vector PKS::GUS was constructed in such a way that the ATG of the PKS gene was in the frame with the β-glucuronidase (GUS) coding region. Using a combinatorial transient GUS expression system in Nicotiana benthamania leaves, it was shown that heterologous TFs such as HlWRKY1, HlMYB3, HlWDR1 and HlbZIP1 from Humulus lupulus significantly activated PKS promoter. Moreover, Tombusvirus p19 core protein, which is known for silencing suppressor functions, acted in our combinatorial transient expression system as an enhancer of PKS promoter activity along with hop TFs. Our analyses suggested the involvement of the hop derived TFs (HlWRKY1, HlMYB3, HlWDR1 and HlbZIP1A) and p19 in the activation of PKS gene promoter, which could be used for the genetic manipulation of C. sativa to enhance the cannabinoid production.

Circular RNAs (circRNAs), an emerging class of non-coding RNAs, are abundant in eukaryotic transcriptomes. Seed germination is one of the most important stages in the entire life cycle of plants that can be slowed down or totally restrained by high temperature. Our aim is to identify heat-responsive circRNAs and explore the potential function of circRNAs in tomato seeds at high temperature. Following high-throughput sequencing, 4 164 circRNAs were identified, and 980 circRNAs were shared in the control and high-temperature libraries. Among the 748 circRNAs with high expressions, 73 circRNAs were significantly up-/down- regulated in tomato seeds germinated at high temperature compared to the control. The parental genes of circRNAs existing in seeds only at high temperature were mainly involved in metabolic processes, cellular processes, catalytic activities, and binding based on Gene Ontology analysis. The results suggested that circRNAs were widespread in tomato and were generated from different chromosomes and diverse genomic regions. Some circRNAs in tomato seeds responded to high temperature during germination. This study provides the first genome-wide profile of circRNAs in response to high temperature during tomato seed germination and lays a foundation for studying the potential biological functions of circRNAs responding to heat stress.

A transcription factor gene Arachis hypogaea abscisic acid (ABA) responsive element binding protein 1 (AhAREB1) has been isolated from peanut previously. Here, the function of different domains from AhAREB1 was investigated using construct series containing AhAREB1 full-length and truncated fragments to transform peanut hairy roots and pAhNCED1 (promoter of Arachis hypogaea 9-cis-epoxycarotenoid dioxygenase 1) GUS/Col Arabidopsis thaliana, respectively. The results of real-time quantitative PCR, transient expression, and chromosome immunoprecipitation (ChIP) assay all showed that AhAREB1 negatively regulated the expression of the AhNCED1 gene. β-Glucuronidase (GUS) staining shows that AhAREB1 and the AhAREB1 gene truncated fragment A1 may be bound to ABA responsive element  motifs in the promoter region of AhNCED1 and involved in the negative regulation of the upstream AhNCED1 gene promoter, reflected by the inhibited expression of the AhNCED1 promoter reporter gene and significantly reduced GUS activity in transgenic A. thaliana plants. Furthermore, only the full variant of AhAREB1 and a fragment without a C1 domain had repression activity on the AhNCED1 promoter. On the contrary, the AhAREB1 gene truncated fragments A2 and A3 variant without a C2 domain had no such repression activity. Moreover, the negative regulation of AhNCED1 was detected only when the C2 domain was present suggesting that the C2 domain was required for AhAREB1 activity. Subcellular localization analysis shows that the deletion of conserved domains C1, C2, C3 had no effects on the nuclear localization of AhAREB1. In addition, ChIP analysis indicates that the deletion of domains C1 and C3 significantly affected the binding of the AhAREB1 transcription factor to the AhNCED1 promoter. Taken together, the results indicate that the different N-terminal domains of the AhAREB1 protein, which played different roles in the negative regulations of AhNCED1, were necessary for AhNCED1 transcription.

In order to understand mechanisms of ontogenic resistance to apple scab, we analyzed various aspects of young and old leaves. We have introduced an apple plants cultivation system where in vitro propagated and rooting explants produce a genetically uniform population of apple (Malus domestica cv. Idared) plants. In this work, we demonstrate that apple plants produced in our cultivation system showed susceptibility to Venturia inaequalis, the cause of apple scab disease in young leaves and resistance in old leaves, which is similar to orchard situation. Our analysis shows that the cessation of epidermal cell expansion and shape formation coincided with the onset of ontogenic resistance in older leaves. Formation of specific cuticular lamellar structures did not coincide with ontogenic resistance onset. Further, chemical composition analysis of wax from young susceptible and old resistant leaves did not reveal specific compounds involved in ontogenic resistance. Differences in homogalacturonan content in cell walls in susceptible and resistant cells as well as decreased methylesterification of pectin in resistant leaves suggest that polysaccharide composition of the cell wall may play a role in mycelium growth and nutrition.

Apyrases belong to the ATPase family of enzymes that hydrolyze phosphoanhydride bonds of nucleoside tri- and di-phosphates. These enzymes differ markedly from other phosphohydrolases due to their high specific activity, broad divalent cation requirement, broad nucleotide substrate specificity, and insensitivity to various inhibitors. In the past 30 years, apyrases have been frequently studied in mammals. In comparison, research of apyrases in plants has received little attention, despite the growth of plants being closely related to the apyrases. In this review, we summarize the research of the apyrases in Arabidopsis thaliana and point to the possible future directions of research. Apyrases have seven members found in Arabidopsis thaliana, each with different properties and functions. Currently, the characterization and functions of AtAPY1 and AtAPY2 have been reported, though, to the best of our knowledge, the other apyrase members (AtAPY3 to 7) have not yet been sufficiently described. In this review, we also summarize the progress being made and the difficulties encountered in apyrase research in Arabidopsis thaliana.

Strawberry tree (Arbutus unedo) is a small perennial tree that grows spontaneously in the Mediterranean basin, Ireland, and Portugal. In this work, strawberry tree clones were established in vitro from epicormic shoots obtained from a young tree, an adult tree, and from a seedling. They were propagated by axillary shoot buds proliferation on solid and in liquid media, and also in a modified De Fossard medium with 9 µM benzylaminopurine. The organogenesis from calli obtained from apical leaves of the in vitro grown shoots from the three genotypes was carried out in the same basal liquid medium supplemented with 9 µM thidiazuron. Micropropagation through organogenesis in liquid medium proved to be more efficient than the other tested methods (considering the number of shoots produced), but the shoots were showing hyperhydricity. Shoots were sucessufully rooted on medium with indole-3-butyric acid and acclimatized ex vitro with rates higher than 90 %. Six month-old plants from the most proliferative genotype (AU1) and propagated in vitro by different methods were submitted to drought stress (no watering for 10 d) and several morphological and physiological parameters were evaluated and compared to a control group (watered to 70 % field capacity). No significant differences were found in plant biomass, root length, and plant height, however, slight differences were observed in water potential, net photosynthetic rate, intercellular CO₂ concentration, and stomatal conductance between the plantlets propagated on solid or liquid medium. In general, the responses to drought stress imposed were was similar in plants micropropagated by different propagation methods.

The increasing world population and limited amount of land area appropriate for intensive agriculture necessitate high-yield cultivars. The focus is on the enrichment of existing crops deficient in nutrients, which is also called biofortification. Microelements, vitamins, and fatty acids belong to most important traits being subjected to biofortification. Biofortification strategies can be divided on fertilization-based strategy, which is characterized by direct application of nutrients or plant growth promoting substances on plants, and biotechnological strategy, which involves molecular biology techniques in order to enhance transport, production, and accumulation of nutrients. Recent advances in plant biotechnology, such as genome-editing, clustered regularly interspaced short palindromic repeats (CRISPR)-associated 9, and transcription activator-like effector nuclease, as well as an extensive study of genetic diversity, are acceptable approaches to the development of biofortified crops.

In plants, calcineurin B-like proteins (CBLs) and CBL-interacting protein kinases (CIPKs) regulate Ca2+ signalling and so responses to biotic and abiotic stresses. However, the details of specific CIPKs functions in various stress responses are poorly understood. Here, we report roles of dicot and monocot CIPK2 genes in response to salinity and heavy metals. Arabidopsis thaliana AtCIPK2 was found to be universally expressed in different tissues and organs and furthermore induced by salinity. Overexpression of AtCIPK2 or Tibetan Plateau wild barley (Hordeum spontaneum) HsCIPK2 in Arabidopsis alleviated toxic effects of NaCl and mercury on seed germination and root growth. Similarly, reduced toxic effects of copper and cadmium on seed germination, but not on root growth, were observed in these transgenic lines. Live-cell fluorescence imaging analysis revealed that HsCIPK2 was predominantly distributed in the cytoplasm and nucleus and weakly localized at the plasma membrane (PM), but its PM association was rapidly enhanced upon exposure to high salinity and mercury. These results suggest an involvement of CIPK2 in plant tolerance to salinity and mercury and provide a new insight into physiological functions of CIPKs in plant response to heavy metals.