Adaptation of cucumber seedlings to low temperature stress by reducing nitrate to ammonium during it's transportation.

BACKGROUND: Low temperature severely depresses the uptake, translocation from the root to the shoot, and metabolism of nitrate and ammonium in thermophilic plants such as cucumber (Cucumis sativus). Plant growth is inhibited accordingly. However, the availability of information on the effects of low temperature on nitrogen transport remains limited. RESULTS: Using non-invasive micro-test technology, the net nitrate (NO3-) and ammonium (NH4+) fluxes in the root hair zone and vascular bundles of the primary root, stem, petiole, midrib, lateral vein, and shoot tip of cucumber seedlings under normal temperature (NT; 26 °C) and low temperature (LT; 8 °C) treatment were analyzed. Under LT treatment, the net NO3- flux rate in the root hair zone and vascular bundles of cucumber seedlings decreased, whereas the net NH4+ flux rate in vascular bundles of the midrib, lateral vein, and shoot tip increased. Accordingly, the relative expression of CsNRT1.4a in the petiole and midrib was down-regulated, whereas the expression of CsAMT1.2a-1.2c in the midrib was up-regulated. The results of 15N isotope tracing showed that NO3--N and NH4+-N uptake of the seedlings under LT treatment decreased significantly compared with that under NT treatment, and the concentration and proportion of both NO3--N and NH4+-N distributed in the shoot decreased. Under LT treatment, the actual nitrate reductase activity (NRAact) in the root did not change significantly, whereas NRAact in the stem and petiole increased by 113.2 and 96.2%, respectively. CONCLUSIONS: The higher net NH4+ flux rate in leaves and young tissues may reflect the higher NRAact in the stem and petiole, which may result in a higher proportion of NO3- being reduced to NH4+ during the upward transportation of NO3-. The results contribute to an improved understanding of the mechanism of changes in nitrate transportation in plants in response to low-temperature stress.

The physiological and molecular mechanism of brassinosteroid in response to stress: a review.

The negative effects of environmental stresses, such as low temperature, high temperature, salinity, drought, heavy metal stress, and biotic stress significantly decrease crop productivity. Plant hormones are currently being used to induce stress tolerance in a variety of plants. Brassinosteroids (commonly known as BR) are a group of phytohormones that regulate a wide range of biological processes that lead to tolerance of various stresses in plants. BR stimulate BRASSINAZOLE RESISTANCE 1 (BZR1)/BRI1-EMS SUPPRESSOR 1 (BES1), transcription factors that activate thousands of BR-targeted genes. BR regulate antioxidant enzyme activities, chlorophyll contents, photosynthetic capacity, and carbohydrate metabolism to increase plant growth under stress. Mutants with BR defects have shortened root and shoot developments. Exogenous BR application increases the biosynthesis of endogenous hormones such as indole-3-acetic acid, abscisic acid, jasmonic acid, zeatin riboside, brassinosteroids (BR), and isopentenyl adenosine, and gibberellin (GA) and regulates signal transduction pathways to stimulate stress tolerance. This review will describe advancements in knowledge of BR and their roles in response to different stress conditions in plants.

24-Epibrassinolide Ameliorates Endogenous Hormone Levels to Enhance Low-Temperature Stress Tolerance in Cucumber Seedlings.

Phytohormone biosynthesis and accumulation are essential for plant growth and development and stress responses. Here, we investigated the effects of 24-epibrassinolide (EBR) on physiological and biochemical mechanisms in cucumber leaves under low-temperature stress. The cucumber seedlings were exposed to treatments as follows: NT (normal temperature, 26 °C/18 °C day/night), and three low-temperature (12 °C/8 °C day/night) treatments: CK (low-temperature stress); EBR (low-temperature and 0.1 µM EBR); and BZR (low-temperature and 4 µM BZR, a specific EBR biosynthesis inhibitor). The results indicated that low-temperature stress proportionately decreased cucumber seedling growth and the strong seedling index, chlorophyll (Chl) content, photosynthetic capacity, and antioxidant enzyme activities, while increasing reactive oxygen species (ROS) and malondialdehyde (MDA) contents, hormone levels, and EBR biosynthesis gene expression level. However, EBR treatments significantly enhanced cucumber seedling growth and the strong seedling index, chlorophyll content, photosynthetic capacity, activities of antioxidant enzymes, the cell membrane stability, and endogenous hormones, and upregulated EBR biosynthesis gene expression level, while decreasing ROS and the MDA content. Based on these results, it can be concluded that exogenous EBR regulates endogenous hormones by activating at the transcript level EBR biosynthetic genes, which increases antioxidant enzyme capacity levels and reduces the overproduction of ROS and MDA, protecting chlorophyll and photosynthetic machinery, thus improving cucumber seedling growth.

Grafting-responsive miRNAs in cucumber and pumpkin seedlings identified by high-throughput sequencing at whole genome level.

Grafting is an important agricultural technique widely used for improving growth, yields and tolerance of crops to abiotic and biotic stresses. As one type of endogenous, non-coding small RNAs, microRNAs (miRNAs) regulate development and responsiveness to biotic and abiotic stresses by negatively mediating expression of target genes at the post-transcriptional level. However, there have been few detailed studies to evaluate the role of miRNAs in mediation of grafting-induced physiological processes in plants. Cucumis sativus and Cucurbita moschata are important vegetables worldwide. We constructed eight small RNA libraries from leaves and roots of seedlings that were grafted in the following four ways: (1) hetero-grafting, using cucumber as scion and pumpkin as rootstock; (2) hetero-grafting, with pumpkin as scion and cucumber as rootstock; (3) auto-grafting of cucumbers and (4) auto-grafting of pumpkins. High-throughput sequencing was employed, and more than 120 million raw reads were obtained. We annotated 112 known miRNAs belonging to 40 miRNA families and identified 48 new miRNAs in the eight libraries, and the targets of these known and novel miRNAs were predicted by bioinformatics. Grafting led to changes in expression of most miRNAs and their predicted target genes, suggesting that miRNAs may play significant roles in mediating physiological processes of grafted seedlings by regulating the expression of target genes. The potential role of the grafting-responsive miRNAs in seedling growth and long-distance transport of miRNA was discussed. These results are useful for functional characterization of miRNAs in mediation of grafting-dependent physiological processes.

Low red to far-red light ratio promotes salt tolerance by improving leaf photosynthetic capacity in cucumber.

Soil salinity severely inhibits leaf photosynthesis and limits agricultural production. Red to far-red light ratio (R/FR) affects leaf photosynthesis under salt stress, however, its regulation mechanism is still largely unknown. This study investigated the effects of different R/FR on plant growth, gas exchange parameters, photosynthetic electron transport, Calvin cycle and key gene expression under salt stress. Cucumber seedlings were exposed to four treatments including 0 mM NaCl and R/FR=7 (L7, control), 0 mM NaCl and R/FR=0.7 (L0.7), 80 mM NaCl and R/FR=7 (H7) and 80 mM NaCl and R/FR=0.7 (H0.7) for 9 days in an artificial climate chamber. The results showed that compared to L7 treatment, H7 treatment significantly reduced relative growth rate (RGR), CO2 assimilation rate (P n), maximum photochemical efficiency PSII (F v/F m), most JIP-test parameters and total Rubisco activity, indicating that salt stress severely inhibited photosynthetic electron transport from PSII to PSI and blocked Calvin cycle in cucumber leaves. However, these suppressions were effectively alleviated by low R/FR addition (H0.7 treatment). Compared to H7 treatment, H0.7 treatment significantly increased RGR and P n by 209.09% and 7.59%, respectively, enhanced F v/F m, maximum quantum yield for primary photochemistry (φ Po), quantum yield for electron transport (φ Eo) and total Rubisco activity by 192.31%, 17.6%, 36.84% and 37.08%, respectively, and largely up-regulated expressions of most key genes involved in electron transport and Calvin cycle. In conclusion, low R/FR effectively alleviated the negative effects of salt stress on leaf photosynthesis by accelerating photosynthetic electron transport from PSII to PQ pool and promoting Calvin cycle in cucumber plants. It provides a novel environmentally friendly light-quality regulation technology for high efficiency salt-resistant vegetable production.

Comparative Physiological and Proteomic Analyses Reveal the Mechanisms of Brassinolide-Mediated Tolerance to Calcium Nitrate Stress in Tomato.

Secondary salinization caused by the overaccumulation of calcium nitrate [Ca(NO3)2] in soils due to excessive fertilization has become one of the major handicaps of protected vegetable production. Brassinolide, a bioactive plant steroid hormone, plays an important role in improving abiotic stress tolerance in plants. However, whether and how brassinolide (BR) can alleviate Ca(NO3)2 stress remains elusive. Here, we investigated the effects of exogenous BR on hydroponically grown tomato (Solanum lycopersicum L.) plants under Ca(NO3)2 stress through proteomics combined with physiological studies. Proteomics analysis revealed that Ca(NO3)2 stress affected the accumulation of proteins involved in photosynthesis, stress responses, and antioxidant defense, however, exogenous BR increased the accumulation of proteins involved in chlorophyll metabolism and altered the osmotic stress responses in tomatoes under Ca(NO3)2 stress. Further physiological studies supported the results of proteomics and showed that the exogenous BR-induced alleviation of Ca(NO3)2 stress was associated with the improvement of photosynthetic efficiency, levels of soluble sugars and proteins, chlorophyll contents, and antioxidant enzyme activities, leading to the reduction in the levels of reactive oxygen species and membrane lipid peroxidation, and promotion of the recovery of photosynthetic performance, energy metabolism, and plant growth under Ca(NO3)2 stress. These results show the importance of applying BR in protected agriculture as a means for the effective management of secondary salinization.

Sugars promote graft union development in the heterograft of cucumber onto pumpkin.

The use of heterografts is widely applied for the production of several important commercial crops, but the molecular mechanism of graft union formation remains poorly understood. Here, cucumber grafted onto pumpkin was used to study graft union development, and genome-wide tempo-spatial gene expression at the graft interface was comprehensively investigated. Histological analysis suggested that resumption of the rootstock growth occurred after both phloem and xylem reconnection, and the scion showed evident callus production compared with the rootstock 3 days after grafting. Consistently, transcriptome data revealed specific responses between the scion and rootstock in the expression of genes related to cambium development, the cell cycle, and sugar metabolism during both vascular reconnection and healing, indicating distinct mechanisms. Additionally, lower levels of sugars and significantly changed sugar enzyme activities at the graft junction were observed during vascular reconnection. Next, we found that the healing process of grafted etiolated seedlings was significantly delayed, and graft success, xylem reconnection, and the growth of grafted plants were enhanced by exogenous glucose. This demonstrates that graft union formation requires the correct sugar content. Furthermore, we also found that graft union formation was delayed with a lower energy charge by the target of rapamycin (TOR) inhibitor AZD-8055, and xylem reconnection and the growth of grafted plants were enhanced under AZD-8055 with exogenous glucose treatment. Taken together, our results reveal that sugars play a positive role in graft union formation by promoting the growth of cucumber/pumpkin and provide useful information for understanding graft union healing and the application of heterografting in the future.

Heterotrimeric G-Protein γ Subunit CsGG3.2 Positively Regulates the Expression of CBF Genes and Chilling Tolerance in Cucumber.

Heterotrimeric guanine nucleotide-binding proteins (G proteins) composed of alpha (Gα), beta (Gß), and gamma (Gγ) subunits are central signal transducers mediating the cellular response to multiple stimuli, such as cold, in eukaryotes. Plant Gγ subunits, divided into A, B, and C three structurally distinct types, provide proper cellular localization and functional specificity to the heterotrimer complex. Here, we demonstrate that a type C Gγ subunit CsGG3.2 is involved in the regulation of the CBF regulon and plant tolerance to cold stresses in cucumber (Cucumis sativus L.). We showed that CsGG3.2 transcript abundance was positively induced by cold treatments. Transgenic cucumber plants (T1) constitutively over-expressing CsGG3.2 exhibits tolerance to chilling conditions and increased expression of CBF genes and their regulon. Antioxidative enzymes, i.e., superoxide dismutase, catalase, peroxidase, and glutathione reductase activities increased in cold-stressed transgenic plants. The reactive oxygen species, oxygen free radical and H2O2, production, as well as membrane lipid peroxidation (MDA) production decreased in transgenic plants, suggesting a better antioxidant system to cope the oxidative-damages caused by cold stress. These findings provide evidence for a critical role of CsGG3.2 in mediating cold signal transduction in plant cells.

Cucumber CsBPCs Regulate the Expression of CsABI3 during Seed Germination.

Cucumber seeds with shallow dormancy start to germinate in fruit that are harvested late. ABSCISIC ACID INSENSITIVE3 (ABI3), a transcription factor in the abscisic acid (ABA) signaling pathway, is one of the most important regulators in the transition from late embryogenesis to germination. Our analysis found a candidate cis-regulatory motif for cucumber BASIC PENTACYSTEINE (CsBPC) in the promoter of CsABI3. Yeast one-hybrid and chromatin immunoprecipitation (ChIP) assays showed that CsBPCs bound to the promoter of CsABI3. Examination of ß-glucuronidase (GUS) activity driven by the CsABI3 promoter in transgenic Arabidopsis thaliana plants overexpressing CsBPCs and a Nicotiana benthamiana (tobacco) luciferase assay indicated that CsBPCs inhibited the expression of CsABI3. Transgenic plants overexpressing CsBPCs were constructed to confirm that CsBPCs participates in the control of seed germination. This study of the cucumber BPC-ABI3 pathway will help to explore and characterize the molecular mechanisms underlying seed germination and will provide necessary information for seed conservation in agriculture and forestry.

Gibberellin Is Involved in Inhibition of Cucumber Growth and Nitrogen Uptake at Suboptimal Root-Zone Temperatures.

Suboptimal temperature stress often causes heavy yield losses of vegetables by suppressing plant growth during winter and early spring. Gibberellin acid (GA) has been reported to be involved in plant growth and acquisition of mineral nutrients. However, no studies have evaluated the role of GA in the regulation of growth and nutrient acquisition by vegetables under conditions of suboptimal temperatures in greenhouse. Here, we investigated the roles of GA in the regulation of growth and nitrate acquisition of cucumber (Cucumis sativus L.) plants under conditions of short-term suboptimal root-zone temperatures (Tr). Exposure of cucumber seedlings to a Tr of 16°C led to a significant reduction in root growth, and this inhibitory effect was reversed by exogenous application of GA. Expression patterns of several genes encoding key enzymes in GA metabolism were altered by suboptimal Tr treatment, and endogenous GA concentrations in cucumber roots were significantly reduced by exposure of cucumber plants to 16°C Tr, suggesting that inhibition of root growth by suboptimal Tr may result from disruption of endogenous GA homeostasis. To further explore the mechanism underlying the GA-dependent cucumber growth under suboptimal Tr, we studied the effect of suboptimal Tr and GA on nitrate uptake, and found that exposure of cucumber seedlings to 16°C Tr led to a significant reduction in nitrate uptake rate, and exogenous application GA can alleviate the down-regulation by up regulating the expression of genes associated with nitrate uptake. Finally, we demonstrated that N accumulation in cucumber seedlings under suboptimal Tr conditions was improved by exogenous application of GA due probably to both enhanced root growth and nitrate absorption activity. These results indicate that a reduction in endogenous GA concentrations in roots due to down-regulation of GA biosynthesis at transcriptional level may be a key event to underpin the suboptimal Tr-induced inhibition of root growth and nitrate uptake. These findings may have important practical implications in effective mitigation of suboptimal temperature-induced vegetable loss under greenhouse conditions.

The physiological and molecular mechanism of brassinosteroid in response to stress: a review

The negative effects of environmental stresses, such as low temperature, high temperature, salinity, drought, heavy metal stress, and biotic stress significantly decrease crop productivity. Plant hormones are currently being used to induce stress tolerance in a variety of plants. Brassinosteroids (commonly known as BR) are a group of phytohormones that regulate a wide range of biological processes that lead to tolerance of various stresses in plants. BR stimulate BRASSINAZOLE RESISTANCE 1 (BZR1)/BRI1-EMS SUPPRESSOR 1 (BES1), transcription factors that activate thousands of BR-targeted genes. BR regulate antioxidant enzyme activities, chlorophyll contents, photosynthetic capacity, and carbohydrate metabolism to increase plant growth under stress. Mutants with BR defects have shortened root and shoot developments. Exogenous BR application increases the biosynthesis of endogenous hormones such as indole-3-acetic acid, abscisic acid, jasmonic acid, zeatin riboside, brassinosteroids (BR), and isopentenyl adenosine, and gibberellin (GA) and regulates signal transduction pathways to stimulate stress tolerance. This review will describe advancements in knowledge of BR and their roles in response to different stress conditions in plants.