GC-MS, alpha-amylase, and alpha-glucosidase inhibition and molecular docking analysis of selected phytoconstituents of small wild date palm fruit (Phoenix pusilla).

Phoenix pusilla (Arecaceae), commonly known as "small wild date palm", is regarded as one of the underutilized fruit crops in South India. Methanol extract of P. pusilla ripened fruits (PPRF) was analyzed for in vitro porcine pancreatic alpha-amylase (PPAA) and rat small intestine alpha-glucosidase (RIAG) inhibition activities, and through gas chromatography-mass spectrometry (GC-MS) analysis. The GC-MS analysis showed the presence of 25 phytoconstituents from PPRF which was further assessed on the docking behavior of five targeted enzymes diabetes mellitus (DM) namely (i) human aldose reductase, (ii) protein tyrosine phosphatase 1B, (iii) pancreatic alpha-amylase, (iv) peroxisome proliferator-activated receptor gamma, and (v) dipeptidyl peptidase IV by using the AutoDock Vina method. In addition to this physicochemical, bioactivity score, absorption, distribution, metabolism, excretion, and toxicity (ADMET) analysis was performed using the Molinspiration and pkCSM free online servers. Methanolic extract of PPRF showed 50% inhibition concentration (IC50) at 69.86 and 72.60 µg/mL levels against PPAA and RIAG enzymes activities, respectively. Interestingly in the present study, GC-MS analysis showed the presence of 25 phytoconstituents from PPRF. Physicochemical analysis of PPRF has exhibited that 13 ligands have complied well with Lipinski's Rule of Five (RoF). With regard to ADMET analysis, one ligand (9,12-octadecadienoic acid [Z,Z]) has predicated to possess both the hepatotoxicity (HT) and skin sensitization (SS) effect. The docking studies showed that 1-formyl-2,5-dimethoxy-6,9,10-trimethyl-anthracene exhibited the maximum atomic contact energy (ACE) for all the five target enzymes of DM. Thus, the current study suggested that the methanolic extract of PPRF and its phytoconstituents could be considered as potent antidiabetic agents.

Enhanced H2S biogenesis followed by its postharvest application retarded senescence development by promoting multiple antioxidant protection systems in button mushroom during cold storage.

Rapid postharvest quality deterioration of button mushroom as fruit body surface browning brings about senescence development and limits its distribution potential and postharvest storage. In this investigation, 0.5 mM of NaHS as the optimum concentration for H2S fumigation was applied to retain the quality of Agaricus bisporus mushrooms concerning some qualitative and biochemical attributes evaluation throughout 15 storage-day at 4°C and 80-90% relative humidity. In H2S fumigated mushrooms, pileus browning index, weight loss and softening decreased, concomitant with higher cell membrane stability as revealed by subsidiary electrolyte leakage, malondialdehyde (MDA) and H2O2 contents compared to the control during the cold storage period. H2S fumigation boosted total phenolics, as presented by the enhanced phenylalanine ammonia-lyase (PAL) activity and total antioxidant scavenging activity, while polyphenol oxidase (PPO) activity diminished. Moreover, in H2S fumigated mushrooms not only peroxidase (POD), catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GR) and glutathione peroxidase (GPx) activities but also ascorbic acid and glutathione (GSH) contents increased, even though glutathione (GSSG) content declined. The raised endogenous H2S level prompted by greater cystathionine ß-synthase (CBS), cystathionine ?-lyase (CSE), cysteine synthase (CS), L-cysteine desulfhydrases (LCD) and D-cysteine desulfhydrases (DCD) enzymes activities until 10d in fumigated mushrooms. In general, the increase in endogenous H2S biogenesis promoted by H2S fumigation in button mushrooms resulted in retarding senescence development, maintaining redox balance by boosting multiple enzymatic and non-enzymatic antioxidants defense parameters.

TEMPRANILLO homologs in apple regulate flowering time in the woodland strawberry Fragaria vesca.

The long juvenile period of fruit trees makes their breeding costly and time-consuming. Therefore, flowering time engineering and shortening the juvenile phase have become a breeding priority for the genetic improvement of fruit tree crops. Many economically valuable fruit trees belong to the Rosaceae family including apples and strawberries. TEMPRANILLO (TEM) acts as a key player in flowering time control through inhibiting FT function. Two genes with high sequence similarity with the Arabidopsis TEM genes were isolated from apple (Malus domestica). Due to the complexity of carrying out functional studies in apple, we characterized their function in woodland strawberry as well as their expression in apple. The expression of MdTEM genes in apple tissues from juvenile plants was dramatically higher than that in the tissues from adult trees. In woodland strawberry, the overexpression of MdTEM genes down-regulated FvFT1, FvGA3OX1, and FvGA3OX2 genes in strawberry. The MdTEM-overexpressing lines exhibited delayed flowering, in terms of days to flowering and the number of leaves at flowering. While, RNAi-mediated silencing of TEM resulted in five days earlier flowering, with a lower number of leaves, a higher trichome density, and in some cases, caused in vitro flowering. According to these results and in silico analyses, it can be concluded that MdTEM1 and MdTEM2 can be considered as orthologs of FvTEM and probably AtTEM genes, which play an important role in regulating the juvenile phase and flowering time through regulating FT and GA biosynthetic pathway.

Characterization of Three SEPALLATA-Like MADS-Box Genes Associated With Floral Development in Paphiopedilum henryanum (Orchidaceae).

Paphiopedilum (Orchidaceae) is one of the world's most popular orchids that is found in tropical and subtropical forests and has an enormous ornamental value. SEPALLATA-like (SEP-like) MADS-box genes are responsible for floral organ specification. In this study, three SEP-like MADS-box genes, PhSEP1, PhSEP2, and PhSEP3, were identified in Paphiopedilum henryanum. These genes were 732-916 bp, with conserved SEPI and SEPII motifs. Phylogenetic analysis revealed that PhSEP genes were evolutionarily closer to the core eudicot SEP3 lineage, whereas none of them belonged to core eudicot SEP1/2/4 clades. PhSEP genes displayed non-ubiquitous expression, which was detectable across all floral organs at all developmental stages of the flower buds. Furthermore, subcellular localization experiments revealed the localization of PhSEP proteins in the nucleus. Yeast two-hybrid assays revealed no self-activation of PhSEPs. The protein-protein interactions revealed that PhSEPs possibly interact with B-class DEFICIENS-like and E-class MADS-box proteins. Our study suggests that the three SEP-like genes may play key roles in flower development in P. henryanum, which will improve our understanding of the roles of the SEP-like MADS-box gene family and provide crucial insights into the mechanisms underlying floral development in orchids.

Exogenous methyl jasmonate enhances phytochemicals and delays senescence in harvested strawberries by modulating GABA shunt pathway.

In order to illustrate the relationship between methyl jasmonate (MeJA) and gamma aminobutyric acid (GABA) in maintaining the quality and improving the postharvest life of strawberry fruit, the harvested fruit were treated with MeJA for 16 h at 20 °C and stored at 3 ± 0.5 °C for 12 days. MeJA enhanced the expression levels of GABA shunt pathway-related genes, including glutamate decarboxylase, GABA transaminase, and succinic semialdehyde dehydrogenase, leading to an increase in GABA accumulation. Treated fruit showed higher levels of total acids, anthocyanins, total phenolics, antioxidants, and phenylalanine ammonia-lyase activity, and lower levels of cell membrane deterioration, total soluble solids, polyphenol oxidase activity and decay incidence rate. The results suggest that the positive effects of MeJA in extending the fruit postharvest life, enhancing phytochemical compounds, and decreasing the decay incidence rate may be due to the effects on motivating GABA shunt pathway and PAL enzyme activity.

CRISPR/Cas9-mediated P-CR domain-specific engineering of CESA4 heterodimerization capacity alters cell wall architecture and improves saccharification efficiency in poplar.

Cellulose is the most abundant unique biopolymer in nature with widespread applications in bioenergy and high-value bioproducts. The large transmembrane-localized cellulose synthase (CESA) complexes (CSCs) play a pivotal role in the biosynthesis and orientation of the para-crystalline cellulose microfibrils during secondary cell wall (SCW) deposition. However, the hub CESA subunit with high potential homo/heterodimerization capacity and its functional effects on cell wall architecture, cellulose crystallinity, and saccharification efficiency remains unclear. Here, we reported the highly potent binding site containing four residues of Pro435, Trp436, Pro437, and Gly438 in the plant-conserved region (P-CR) of PalCESA4 subunit, which are involved in the CESA4-CESA8 heterodimerization. The CRISPR/Cas9-knockout mutagenesis in the predicted binding site results in physiological abnormalities, stunt growth, and deficient roots. The homozygous double substitution of W436Q and P437S and heterozygous double deletions of W436 and P437 residues potentially reduced CESA4-binding affinity resulting in normal roots, 1.5-2-fold higher plant growth and cell wall regeneration rates, 1.7-fold thinner cell wall, high hemicellulose content, 37%-67% decrease in cellulose content, high cellulose DP, 25%-37% decrease in cellulose crystallinity, and 50% increase in saccharification efficiency. The heterozygous deletion of W436 increases about 2-fold CESA4 homo/heterodimerization capacity led to the 50% decrease in plant growth and increase in cell walls thickness, cellulose content (33%), cellulose DP (20%), and CrI (8%). Our findings provide a strategy for introducing commercial CRISPR/Cas9-mediated bioengineered poplars with promising cellulose applications. We anticipate our results could create an engineering revolution in bioenergy and cellulose-based nanomaterial technologies.

Proteomic, biochemical, and anatomical influences of nanographene oxide on soybean (Glycine max).

Nano-graphene oxide (NGO) is an engineered nanostructure that is used in various fields including biology, chemistry, medicine, and environmental protection. This kind of highly used nanomaterial (NM) is being released and accumulated gradually in nature and can have some adverse influences on living organisms including plants. Soybean as a cultivated plant with a high importance in food industry, but sensitive to stresses, was chosen in the present study to be examined in terms of proteomic, biochemical, and anatomical properties under the NGO stress. Accordingly, a 2-dimensional gel electrophoresis (2-DE) approach was adopted for proteomic analysis of the NGO treated soybean roots, where significant changes were observed in the abundance of 48 proteins. MALDI TOF/TOF analysis revealed the upregulation of the proteins involved in the redox regulation in plants. Furthermore, anatomical examination of soybean roots under light microscopy showed that the NGO could enter into the root epidermis through the apoplastic pathway and accumulated in some parts of the root. With increasing NGO concentration, the diameter of the vascular apertures increased and then decreased at higher concentrations. To evaluate the toxicity of NGO, some of the growth parameters including fresh and dry weight, and height of the shoots, as well as some stress-related biochemical properties such as H2O2 production, antioxidant enzymes activity, and phenolics and flavonoids contents were measured. The results indicated that NGO could cause an oxidative stress, which can be considered a toxic effect evoking antioxidative and detoxification mechanisms in soybean.

Improvement of Postharvest Quality of Plum (Prunus domestica L.) Using Polysaccharide-Based Edible Coatings.

Polysaccharide-based edible coatings are served as an attractive preservation method for postharvest maintenance of most fruits. The current study examined the effect of carboxymethylcellulose (CMC)- and pectin (Pec)- based edible coatings on titratable acidity (TA), firmness; vitamin C (vit C); total soluble solids (TSS); pH; total phenolics; anthocyanin and flavonoid contents; total antioxidant capacity (based on 1,1-Diphenyl-2-picryl-hydrazyl hydrate (DPPH)); the activities of peroxidase (POD), polyphenol oxidase (PPO) and polygalacturonase (PG) enzymes; and weight loss during cold storage. The results showed that each coating and their combinations caused positive effects in all measured parameters except weight loss. The applied coatings preserved firmness and improved total phenols, anthocyanin and flavonoid contents, antioxidant capacity and POD activity. In addition, TSS decreased and pH values remained more or less stable with the coating application. The coatings delayed TA and vitamin C loss, and decreased enzymatic activities such as PPO and PG. It could be stated that CMC at 1% and Pec at 1.5% separately demonstrated the best results for most of the measured parameters; and 0.5% Pec + 1.5% CMC could be considered as the best combination. In conclusion, application of CMC, Pec, or their combinations would be considered as an interesting approach to improve postharvest quality characteristics of plum fruit.

Pectin-based edible coating preserves antioxidative capacity of plum fruit during shelf life.

Exploiting safer methods for fruit preservation such as application of edible coatings can improve shelf life, valuable characteristics, and antioxidative capacity. The current study aimed to investigate the effect of a pectin-based edible coating on antioxidative capacity of plum fruit during shelf life (19 ± 2 ℃ and 65% relative humidity for eight days). To do this, three solutions (0.5, 1, and 1.5%) of pectin, plasticized by glycerol (0.3% w/v), were applied on plum fruit and compared to a control treated with only distilled water. Ascorbic acid, total phenolics, anthocyanin and flavonoid contents, total antioxidative capacity based on 1,1-diphenyl-2-picryl-hydrazyl hydrate method, peroxidase (as an antioxidant enzyme), and polyphenol oxidase (as an oxidant enzyme) activities were recorded during this period. The results demonstrated that pectin-based edible coating was significantly effective on maintaining ascorbic acid, anthocyanin and flavonoid contents, and antioxidative capacity in plum fruits (P ≤ 0.01). The activities of enzymes were significantly affected by the coatings; peroxidase activity increased and polyphenol oxidase activity decreased (P ≤ 0.01). All pectin concentrations significantly caused higher ascorbic acid and anthocyanin contents, antioxidative capacity, and peroxidase activity but a lower polyphenol oxidase activity than the control; however, just 1 and 1.5% concentrations were effective in terms of total phenolic compounds and flavonoid content, respectively, and the other concentrations acted the same as the control. In general, the coating constituted from 1.5% pectin showed the best results for most measured parameters. Considering the influences of pectin-based edible coating on antioxidative characteristics of plum fruits, its application can be potentially regarded as a favorable method to enhance nutritional value of fruits.

Sodium Chloride Induced Stress Responses of Antioxidative Activities in Leaves and Roots of Pistachio Rootstock.

Salinity substantially affects plant growth and crop productivity worldwide. Plants adopt several biochemical mechanisms including regulation of antioxidant biosynthesis to protect themselves against the toxic effects induced by the stress. One-year-old pistachio rootstock exhibiting different degrees of salinity tolerance were subjected to sodium chloride induced stress to identify genetic diversity among cultivated pistachio rootstock for their antioxidant responses, and to determine the correlation of these enzymes to salinity stress. Leaves and roots were harvested following NaCl-induced stress. The results showed that a higher concentration of NaCl treatment induced oxidative stress in the leaf tissue and to a lesser extent in the roots. Both tissues showed an increase in ascorbate peroxidase, superoxide dismutase, catalase, glutathione reductase, peroxidase, and malondialdehyde. Responses of antioxidant enzymes were cultivar dependent, as well as temporal and dependent on the salinity level. Linear and quadratic regression model analysis revealed significant correlation of enzyme activities to salinity treatment in both tissues. The variation in salinity tolerance reflected their capabilities in orchestrating antioxidant enzymes at the roots and harmonized across the cell membranes of the leaves. This study provides a better understanding of root and leaf coordination in regulating the antioxidant enzymes to NaCl induced oxidative stress.

NPR1 as a transgenic crop protection strategy in horticultural species.

The NPR1 (NONEXPRESSOR OF PATHOGENESIS RELATED GENES1) gene has a central role in the long-lasting, broad-spectrum defense response known as systemic acquired resistance (SAR). When overexpressed in a transgenic context in Arabidopsis thaliana, this gene enhances resistance to a number of biotic and abiotic stresses. Its position as a key regulator of defense across diverse plant species makes NPR1 a strong candidate gene for genetic engineering disease and stress tolerance into other crops. High-value horticultural crops face many new challenges from pests and pathogens, and their emergence exceeds the pace of traditional breeding, making the application of NPR1-based strategies potentially useful in fruit and vegetable crops. However, plants overexpressing NPR1 occasionally present detrimental morphological traits that make its application less attractive. The practical utility of NPR-based approaches will be a balance of resistance gains versus other losses. In this review, we summarize the progress on the understanding of NPR1-centered applications in horticultural and other crop plants. We also discuss the effect of the ectopic expression of the A. thaliana NPR1 gene and its orthologs in crop plants and outline the future challenges of using NPR1 in agricultural applications.

Ion homeostasis, osmoregulation, and physiological changes in the roots and leaves of pistachio rootstocks in response to salinity.

Pistachio, one of the important tree nuts, is cultivated in arid and semi-arid regions where salinity is the most common abiotic stress encountered by this tree. However, the mechanisms underlying salinity tolerance in this plant are not well understood. In the present study, five 1-year-old pistachio rootstocks (namely Akbari, Badami, Ghazvini, Kale-Ghouchi, and UCB-1) were treated with four saline water regimes (control, 8, 12, and 16 dS m-1) for 100 days. At high salinity level, all rootstocks showed decreased relative water content (RWC), total chlorophyll content (TCHC), and carotenoids in the leaf, while ascorbic acid (AsA) and total soluble proteins (TSP) were reduced in both leaf and root organs. In addition, the total phenolic compounds (TPC), proline, glycine betaine, total soluble carbohydrate (TSC), and H2O2 content increased under salinity stress in all studied rootstocks. Three different ion exclusion strategies were observed in the studied rootstocks: (i) Na+ exclusion in UCB-1, because most of its Na+ is retained in the roots; (ii) Cl- exclusion in Badami, in which most of its Cl- remained in the roots; and (iii) similar concentrations of Na+ and Cl- were observed in the leaves and roots of Ghazvini, Akbari, and Kale-Ghouchi. Transport capacity (ST value) of K+ over Na+ from the roots to the leaves was more observable in UCB-1 and Ghazvini. Overall, the root system cooperated more effectively in UCB-1 and Badami for retaining and detoxifying an excessive amount of Na+ and Cl-. The results presented here provide important inputs to better understand the salt tolerance mechanism in a tree species for developing more salt-tolerant genotypes. Based on the results obtained here, the studied rootstocks from tolerant to susceptible are arranged as follows: UCB-1 > Badami > Ghazvini > Kale-Ghouchi > Akbari.

Salicylic acid affects the expression of VvCBF4 gene in grapes subjected to low temperature.

The present study investigates the effects of exogenous salicylic acid (SA) on the expression of Vitis vinifera C-repeat binding factor 4 (VvCBF4) gene under low-temperature conditions in an Iranian Vitis viniferea L. 'Sultanina'. The experiment was conducted as a factorial experiment based on a completely randomized design with four replications. 100 µmol/L SA (0, 1, 6 and 12 h before applying cold stress) in temperatures of 1 ± 0.5 °C (for 1, 3, 6 and 12 h) and 22 °C (as control) were applied. The highest expression was observed in plants treated 6 h before sampling. By increasing the duration of low temperature, the expression of VvCBF4 increased. Increasing the duration of cold stress to 6 h in 1 °C increased the expression of VvCBF4 to 24.3 fold. Exogenous application of SA and cold stress treatments increased the expression of VvCBF4. In conclusion, exogenous application of SA in cold stress, increased the expression of VvCBF4 depending on treating time before cold stress. The highest VvCBF4 expression was observed in plants treated 6 h before sampling and increasing the time decreased the expression. By increasing the expression of VvCBF4 the tolerance of plant to cold stress increased.

Green-tissue-specific, C(4)-PEPC-promoter-driven expression of Cry1Ab makes transgenic potato plants resistant to tuber moth (Phthorimaea operculella, Zeller).

An important strategy for obtaining a safer transgenic plant may be the use of a spatial- or tissue-specific promoter, instead of a constitutive one. In this study, we have used a light-inducible maize PEPC promoter to regulate the cry1Ab gene, aiming to produce transgenic potatoes that are resistant to potato tuber moth (PTM) (Phthorimaea operculella, Zeller). Out of 60 regenerated lines having normal phenotypes, 55 lines were PCR-positive for both the cry1Ab and nptII genes. Southern analysis on three selected putative transgenic lines revealed that they have only a single intact copy of the cry1Ab gene. An investigation of the Cry1Ab protein in the leaves and light-exposed (LE) tubers of the transgenic lines demonstrated the presence of the protein in the foliage and green tubers but not in the light-not exposed (LNE) tubers. A bioassay analysis of excised leaves of nine randomly selected lines showed that eight lines had 100% PTM larval mortality. Confirming results were obtained in six selected lines using the whole plant bioassay in the greenhouse. LE transgenic tubers also exhibited 100% larval mortality; however, the levels of damage to the LNE transgenic tubers were high and statistically the same as those incurred by the non-transgenic ones. Based on the results, we believe that this spatial expression of Cry1Ab using the light-inducible PEPC promoter can control PTM infestation in the field and significantly reduce pollution transmission to storage potatoes.