Doctoral Dissertation Hormonal regulation of drought stress responses and tolerance in Brassica napus L Department of Animal Science and Bioindustry Graduate School, Chonnam National University La Van[.]
GENERAL INTRODUCTION
Brassica species and drought stress
Brassica species, including Brassica napus, are vital vegetables that provide essential oils and proteins for both humans and animals As a significant agro-economic crop, rapeseed oil serves as a source of polyunsaturated fatty acids, making it a healthy dietary ingredient Additionally, canola oil is valued in animal feeds due to its high protein content, approximately 50% However, like other field crops, oilseed rape is particularly vulnerable to environmental stress (Zhu et al., 2016; Elferjani and Soolanayakanahally, 2018).
Drought is a significant environmental stress that adversely affects the growth, development, and productivity of crops, particularly canola This stress impacts the flowering and grain filling stages, leading to substantial reductions in both seed yield and quality Brassica vegetables, which are composed of over 85% water, are particularly sensitive to water deficits and excesses, affecting their overall yield and quality Recent advancements in engineering have enabled the development of crops that can withstand stress, highlighting the importance of understanding the molecular defense mechanisms for enhancing stress tolerance in Brassica napus.
ROS is a primary stress signal for metabolism and transduction signaling 3 1 ROS generation and metabolism
Reactive oxygen species (ROS) are highly reactive forms of oxygen molecules, including hydroxyl radicals (HO-), superoxide (O2-), hydrogen peroxide (H2O2), and singlet oxygen (1O2) These ROS are produced by the plasma membrane NADPH oxidase, known as Respiratory Burst Oxidase Homologues (RBOHs) in plants, as well as by peroxidases and oxidases present in the apoplast (Wrzaczek et al., 2013; Waszczak et al.).
2018) It has been known the cell organelles consist of the chloroplast, mitochondria,
4 and peroxisomes are major sources of intracellular ROS production in plant cells through photosynthesis, respiration, and photorespiration (Figure 1.1)
The main compartment for hydrogen peroxide (H2O2) production in photosynthetic cells involves several key enzymes, including glycolate oxidase (GO), peroxidase (POX), and xanthine oxidase (XO) Additionally, crucial metabolic intermediates such as 3-phosphoglycerate (3PGA) and ribulose 1,5-bisphosphate (RuBP) are involved in this process, with ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO) playing a significant role Superoxide dismutase (SOD) is also essential in managing reactive oxygen species within the cell, highlighting the intricate network of enzymatic functions that contribute to H2O2 production in photosynthesis (Mhamdi et al., 2010b).
Reactive oxygen species (ROS) play a crucial role in helping plants acclimate to various stresses by altering their internal and external metabolism In chloroplasts, ROS are key players in signal transduction, facilitating interactions between plants and their environment The formation of hydrogen peroxide in chloroplasts triggers changes in the antioxidant system, serving as signals that communicate environmental and metabolic changes to the nucleus through chloroplast retrograde signaling (Estavillo et al., 2013; Noctor and Foyer, 2016; Waszczak et al., 2018).
1.2.2 ROS role in transmitting signal
Hydrogen peroxide (H2O2) has been recognized as a crucial signaling molecule, acting as a second messenger in various biological processes (Rejeb et al., 2014; Sies, 2017; Winterbourn, 2017) The role of reactive oxygen species (ROS) in signal transduction highlights the importance of regulatory networks that balance ROS generation and scavenging, thereby controlling ROS responses and subsequent downstream processes (Mittler, 2004) Research indicates that ROS generation enhances antioxidant enzyme activity in response to abiotic stresses across different plant species (Lee et al., 2008; Mhamdi et al., 2010b) Central to this regulatory mechanism is the production of ROS.
Recent studies indicate that calcium ions (Ca 2+) play a crucial role in stress responses by interacting with reactive oxygen species (ROS) signaling, mitogen-activated protein kinases (MAPKs), and hormones An increase in cytosolic Ca 2+ serves as an upstream signal that regulates ROS production through the modification of NADPH oxidase by calcium-dependent protein kinases (CDPKs) Additionally, Ca 2+ is involved in downstream ROS signaling and activates antioxidant defenses, while MAPKs are linked to hormone production and signaling In stressed plants, the activation of both ROS, primarily H2O2, and phytohormone signaling is essential for acclimating to stress conditions.
Proline is an elicitor in plants response to drought stress
1.3.1.Proline accumulation in stress response
Proline is an amino acid that plays a crucial role in the response to various abiotic stresses, including drought, salt, oxidative, and pathogen stresses It is recognized as a compatible osmolyte that enhances drought stress tolerance Additionally, proline acts as an osmoprotectant, helping to stabilize membranes and detoxify tissues affected by excess stress.
Proline accumulation in plants during stress is regulated by the balance between its synthesis and degradation Under water stress, proline synthesis is enhanced while its breakdown is inhibited Conversely, re-watering reverses this regulation, leading to increased degradation of proline.
Figure 1.2 Model for proline synthesis pathway in plants There are two ways activates from ornithine d-aminotransferase (OAT) and glutamate (Glu)
Proline is primarily synthesized from glutamate through the glutamate and ornithine pathways, particularly during abiotic stress, involving key enzymes such as P5CS and OAT in the cytosol and chloroplasts Its catabolism occurs in mitochondria, where proline dehydrogenase and P5C dehydrogenase convert proline to glutamate Additionally, proline transport plays a significant role in its accumulation under stress conditions, notably observed in the phloem sap of water-stressed plants.
1.3.2.Proline metabolism essential for stress response and tolerance
Research indicates that proline plays a crucial role in mechanisms of resistance beyond osmotic adjustments, involving specific morphological and physiological modifications (Deuschle et al., 2004; Chen et al., 2011; Lee et al., 2013) Furthermore, both exogenous application and endogenous overproduction of proline have demonstrated a significant delay in cell death.
2004) Available evidence indicates that high proline intracellular induces cell death by increasing ROS production (Chen et al., 2011) The generation of ROS might be
The proline degradation pathway, specifically the P5C–proline cycle, plays a crucial role in enhancing the hypersensitive response in Arabidopsis mutants overexpressing PDH or P5CDH This process increases the likelihood of electrons being produced from proline oxidation to glutamate, thereby elevating the rate of electron transfer.
Proline degradation plays a crucial role in regulating cellular ROS homeostasis, which can impact endogenous signaling pathways (Rejeb et al., 2014; Szabados and Savoure, 2010) Numerous reviews highlight that proline accumulation in stressed plants functions as a signaling molecule, influencing various metabolic regulatory pathways (Szabados).
Proline plays a crucial role in plant response to abiotic stress by interacting with reactive oxygen species (ROS), antioxidants, redox balance, and hormonal signals Research has shown that proline metabolism is linked to changes in ROS levels and the abscisic acid (ABA) signaling pathway, which collectively enhance stress tolerance in plants.
Arabidopsis mutant and transgenic plants, Deuschle et al (2004) reported that seem to be altered SA level or SA signaling might be related to oxidative stress-induced by proline toxicity.
Redox balance: A tools of plant defense against drought stress
Reactive oxygen species (ROS) stimulation is often linked to a highly activated antioxidant scavenging system In plant cells, glutathione (GSH) and ascorbic acid (AsA) serve as the primary nonenzymatic antioxidants, crucial for maintaining intracellular redox homeostasis GSH, a thiol, plays a vital role in stabilizing Calvin cycle enzymes and ensuring that AsA remains in its reduced form within chloroplasts.
Maintaining high ratios of glutathione (GSH) to oxidized glutathione (GSSG) is essential for redox balance, particularly due to the role of ascorbate (AsA) in the AsA-GSH cycle This cycle is crucial for the metabolism of hydrogen peroxide (H2O2) during oxidative stress H2O2 reduction is connected to NADPH oxidation through the interplay of ascorbate and glutathione Additionally, ascorbate peroxidase (APX) activity can facilitate NADPH oxidation independently of GSH through the action of monodehydroascorbate reductase (MDHAR) Furthermore, GSH can chemically reduce dehydroascorbate (DHA) through an enzymatic link with the AsA-GSH pool.
The enzyme DHAR catalyzes the conversion of ascorbate, highlighting the critical role of GSH-dependent ascorbate pools in regulating stress tolerance The ascorbate-glutathione (AsA-GSH) cycle is essential for detoxifying reactive oxygen species, underscoring its significance in various physiological processes.
H2O2 and protects from oxidative stress.
Plant responses to drought stress: A matter of hormones regulation
Plants under stress activate H2O2 and phytohormone signaling pathways (Xia et al., 2015; Choudhury et al., 2017) During water stress, abscisic acid (ABA) plays a crucial role in regulating plant responses and interacts with other hormones such as jasmonic acid (JA), salicylic acid (SA), cytokinins (CK), gibberellic acid (GA), and indole-3-acetic acid (IAA) (Yang et al., 2003; Yasuda et al., 2008; Zhao et al., 2014; Muñoz-Espinoza et al., 2015; Huang et al., 2018; Wang et al., 2018).
Abscisic acid (ABA) is a crucial stress phytohormone that rapidly accumulates in plants, triggering the expression of stress-responsive genes and facilitating physiological adaptations to drought stress Under stress conditions, plants activate ABA-dependent responses through the perception of ABA by pyrabactin resistance 1 (PYR1) and PYR1-like (PYL) proteins These PYLs inhibit clade A protein phosphatase 2Cs (PP2Cs) in an ABA-dependent manner, leading to the activation of sucrose non-fermenting 1-related protein kinases (SnRK2s) Activated SnRK2s play a vital role in regulating ABA-responsive gene expression by phosphorylating transcription factors, such as ABA-responsive element-binding factors (ABFs), and other substrates involved in various physiological processes Overall, SnRK2s serve as central regulators in the ABA signaling pathway, mediating the activation of ABA-responsive transcription factors and genes essential for physiological responses to stress.
2009; Umezawa, 2009; Kulik et al., 2011; Yoshida et al., 2015) In unstressed plants, the target of rapamycin (TOR) kinase phosphorylates PYLs to prevent activation of stress responses (Wang et al., 2018)
Salicylic acid (SA) is crucial for regulating plant development and responses to both biotic and abiotic stresses It plays a significant role in enhancing basal resistance to abiotic stresses and acts as a key signal in disease resistance mechanisms Research on Arabidopsis mutants has demonstrated that endogenous SA accumulation promotes disease resistance and water-deficit tolerance SA functions within a signaling pathway involving SA-inducible genes, particularly the nonexpressor pathogenesis-related gene 1 (NPR1) and pathogenesis-related (PR) proteins NPR1 serves as the central regulator in the SA signaling pathway, facilitating interactions with TGA2 and TGA3 to activate a range of PR genes through RNA polymerase II Furthermore, SA-induced drought tolerance is linked to the expression of PR genes, such as PR1, PR2, and PR5, which are commonly used as markers for SA-mediated drought resistance.
1.5.3.Antagonism between abscisic acid and salicylic acid pathways
Plants respond to stress by altering hormone levels, particularly abscisic acid (ABA) and salicylic acid (SA), which are crucial for their stress responses Research on mutant plants has shown that ABA enhances basal resistance to abiotic stress, indicating its interaction with SA during stress conditions Antagonistic interactions between these hormones play a significant role in regulating defense responses Additionally, hormone crosstalk, observed in both abiotic and biotic stress, highlights the complex metabolic interactions involved The interplay between ABA and SA in plant defense mechanisms is well-documented, with NPR1 serving as a key component in the SA-ABA antagonism.
Systemic acquired resistance (SAR) signaling downstream of salicylic acid (SA) plays a crucial role in regulating the interaction between SA and abscisic acid (ABA) signaling, as highlighted by Yasuda et al (2008) and Muñoz-Espinoza et al (2015) Research on tomato plants indicates that their adaptation to osmotic stress is influenced by SA concentration, revealing challenges in replicating the precise balance and timing of endogenous hormone levels Mur et al (2006) found that low SA concentrations can enhance hormone synergy, while high concentrations may lead to antagonism Despite the documented reciprocal inhibition, the relationship between SA and ABA is not exclusively antagonistic; for example, SA treatments have been shown to increase ABA and proline levels in barley leaves (Bandurska and Stroinski, 2005; Wang et al., 2018) Nonetheless, the dynamics between SA and ABA signaling during water deficit conditions remain largely unexplored.
Carbon and nitrogen metabolism in stress tolerance
1.6.1.Sucrose as a component of carbon source
Sugars, particularly sucrose, play a crucial role in plant development and stress tolerance regulation (Zheng et al., 2010; Ma et al., 2017; Sakr et al., 2018) The accumulation of sugars results from hexose biosynthesis via hexokinase (HXK) phosphorylation (Moore et al., 2003; Ruan, 2014) and sucrose synthesis through sucrose phosphate synthase (Baxter et al., 2003), along with starch degradation and sugar transport (Durand et al., 2018) Sucrose is well-known as a signaling molecule in phloem loading, influencing the partitioning of photo-assimilates between source and sink (Gong et al., 2015; Ruan, 2014) The sucrose sensing pathway involves calcium and calcium-dependent protein kinases (CDPKs) that transmit sucrose signals and interact with bZIP transcription factors (Toyota et al., 2018; Sakr et al., 2018) Additionally, the SnRK1/2 and PP2C phosphatase interaction is significant in the ABA signaling pathway, with shared downstream targets for metabolic signaling Importantly, some gene responses linked to the SnRK1 pathway may operate independently of HXK1 signaling, highlighting the complexities of sucrose sensors and signaling in hormone regulation.
The ongoing research indicates that factors such as sucrose synthetase and sucrose-degrading enzymes (Nguyen et al., 2015), along with the BZR1-BAM transcription factor, which interacts with a non-catalytic amylase-like domain (Soyk et al., 2014), may play a significant role in this regulatory mechanism.
1.6.2 Proline integrates nitrogen assimilation pathway
Proline accumulation plays a crucial role in abiotic stress tolerance by serving as a source of nitrogen and carbon metabolism The synthesis of proline primarily relies on glutamate, which is considered a key substrate in this process Research indicates that glutamate acts as a significant precursor for proline synthesis and is also involved in ammonium assimilation The availability of glutamate is regulated by nitrogen assimilation, mainly through the enzyme glutamine synthetase (GS), which converts glutamate and ammonium into glutamine, while glutamate synthetase (GOGAT) generates glutamate For instance, nitrogen starvation in wheat seedlings has been shown to significantly reduce the levels of NR, GS, and GOGAT at both transcript and enzyme activity levels Additionally, glutamate dehydrogenase (GDH) in mitochondria can directly produce glutamate from ammonium The relationship between nitrogen assimilation, transport, and proline accumulation in hormone signaling during water deficit conditions remains complex and not fully understood.
Understanding the hormonal network involved in metabolite changes during plant responses to water stress is crucial Deciphering the interactions between hormones, reactive oxygen species (ROS), sucrose, proline, and redox status reveals emerging mechanisms that contribute to drought stress tolerance.
Chapter 1, 2 and 3 to assess the significance of the phytohormones in plant response, as follows;
1) Assess the effect of salicylic acid to plant physiology and biochemicals (proline and redox) response under drought stress
2) To elucidate the interplay between reactive oxygen species, proline, and redox state in hormones antagonistic
3) To elucidate the underlying mechanism SA induces sucrose accumulation contributes to drought tolerance
Chapter 4 was to elucidate the interplay between hormonal and hydrogen peroxide regulations in nitrogen metabolism in drought stress response, as described below;
1) To address the significance of SA and H2O2 interplays in proline metabolism-mediated nitrogen assimilation
Table 1 List of the primers used in this study
Name Accession No Oligonucleotide sequence of primers
R: 5′- TGTTCCCATTGCCCTGTG -3´ BrP5CSA XM-009143589 F: 5′- GGTCATGCTGATGGAATCTG -3´
R: 5′- GCATTACAGGCTGCTGGATA -3´ BrP5CSB XM-009118014 F: 5´- TGGACAGAGCAGTCTCATGG -3´
R: 5´- GCACGCATGAAATCAGAGAA -3´ BnCu/ZnSOD AY970822 F: 5´- TGCTAATCGTCATGCTGGAG -3´
R: 5´- AAGCAGCTTGTCATCCGAGT -3´ BnCHLG XM_013788949.1 F: 5´- CTACGAACTCGTCAC CAAAG -3´
14 All primers were designed directly from sequences in the public database, F: Forward; R: Reverse
MATERIALS AND METHODS
Plant culture
Chinese cabbage (Brassica campestris var pekinensis) was cultivated in 2-L pots filled with a soil and perlite mixture (70:30, v/v) in a glasshouse The growing conditions included a mean day/night temperature of 25:20 °C, relative humidity of 50-70%, and a photoperiod of 16 hours of light followed by 8 hours of darkness To enhance natural light, an additional 200 μmol m -2 s -1 was provided at the canopy height for 16 hours each day Complete nutrient solutions, containing both macroelements and microelements, were continuously supplied until the plants were ready for harvest (Lee et al., 2013).
Brassica napus (cv Capitol) seeds were sown in bed soil in a tray Plants with the
At the 4-leaf stage, plants were transferred to 2-L pots filled with a soil and perlite mixture (70:30, w/w) and cultivated in a greenhouse A continuous supply of complete nutrient solution was provided (Lee et al., 2015), while natural light was enhanced with metal halide lamps, delivering approximately 200 μmol photons m -2 s -1 at the canopy height for 16 hours daily.
Introduction
Salicylic acid (SA) is a phytohormone that regulates plant defenses against pathogens and abiotic stresses, enhancing protective reactions that boost photosynthesis and CO2 assimilation Its application in water-deficient plants mitigates cell membrane damage by reducing lipid peroxidation and H2O2 accumulation SA also enhances the activities of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), and guaiacol peroxidase (GPOD), while modulating the glutathione (GSH) and ascorbate (AsA) cycles to combat drought stress GSH and AsA serve as key non-enzymatic antioxidants that detoxify H2O2, alleviating oxidative stress during abiotic challenges Furthermore, SA treatment has been shown to significantly increase the levels of AsA and GSH, as well as the expression of genes related to the AsA-GSH cycle, thereby improving stress tolerance in plants.
SA in plant stress resistance has long been known, the physiological mechanism regarding this stress tolerance remains largely unknown
Proline accumulation is a key response in many plants to environmental stressors like drought, serving as a compatible solute that helps maintain turgor pressure and protect cellular structures This process not only enhances plant resistance to adverse conditions but also plays a crucial role in detoxifying ammonia and reactive oxygen species (ROS), while activating antioxidant enzymes such as catalase (CAT).
Research has highlighted the significance of proline metabolism in enhancing stress tolerance, particularly under conditions like salinity and drought (Iqbal et al., 2014) Overexpression of proline biosynthetic enzymes, such as P5C synthase 1 (P5CS1) and P5C reductase (P5CR), leads to increased proline levels and reduced hydrogen peroxide (H2O2) and malondialdehyde (MDA), which are indicators of oxidative stress (Zhu et al., 1998) In contrast, p5cs1 mutants exhibit decreased proline levels and heightened oxidative damage under salt stress (Székely et al., 2008) The enzymes proline dehydrogenase (PDH) and P5C dehydrogenase (P5CDH) facilitate the conversion of proline back to glutamine, with their activities suppressed in photosynthetic tissues, thereby enhancing resistance to salinity and freezing (Nanjo et al., 1999) Recent studies have established a connection between proline metabolism and cellular redox status in drought-stressed plants, where NADPH serves as an electron donor in proline biosynthesis, contributing to a balanced NADPH/NADP+ ratio and reducing singlet oxygen production in the thylakoid membrane (Sharma et al., 2011; Giberti et al., 2014) Additionally, salicylic acid (SA) has been shown to promote proline accumulation by activating its metabolic enzymes (Khan et al., 2013) However, the regulatory mechanisms of proline synthesis and its interaction with SA in the context of drought stress tolerance remain inadequately explored.
This study investigates the role of salicylic acid (SA) pretreatment in maintaining cellular redox homeostasis by regulating proline biosynthesis during drought stress Plants were treated with SA for seven days before being subjected to 14 days of drought stress The research measured reactive oxygen species (ROS) levels, antioxidant enzyme activity, redox status (including GSH and GSSG), pyridine nucleotides, and the processes of proline biosynthesis and degradation.
Experiment design
Approximately 10 weeks later, plants were treated without (control) or with salicylic acid (SA) For the SA treatment, 30 mL of 1.5 mM SA was sprayed to 6
In a study involving 26 plants, a consistent volume of water was administered to 6 control plants over a period of 7 days Subsequently, half of both the control and salicylic acid (SA)-pretreated plants were subjected to drought stress The well-watered control group received daily irrigation of 150 mL, while the drought-stressed plants were given only 30 mL of nutrient solution per pot.
In a drought-stress treatment lasting 14 days, 30 mL of a nutrient solution equivalent to that used for the control pot was applied Following this treatment, leaves were harvested and preserved at -80°C for subsequent analysis Key metrics evaluated included changes in plant morphology, biomass, stress symptoms, and biochemical responses under both salicylic acid (SA) and drought conditions.
Figure 1 Experimental design of salicylic acid treated to Chinese cabbage plants under non-drought and drought stress conditions.
Results
Effect of SA pretreatment on biomass, osmotic potential, and chlorophyll and carotenoid content under drought stress conditions
Drought stress caused visible wilting in Chinese cabbage leaves, although this effect was less pronounced in plants pretreated with salicylic acid (SA) Biomass reduction was significant, with a 28% decrease in drought-stressed plants without SA treatment, compared to a 9.8% decrease in those with SA pretreatment Additionally, drought stress led to a 41.6% reduction in osmotic potential in non-SA pretreated plants, while SA pretreated plants experienced a 12.6% decrease, relative to the control group.
Table 1.1 Changes in the fresh weight of Brassica rapa in the control or salicylic acid pretreated plants under well–watered or drought stressed conditions
Values are presented as means ± SE for n = 3 Values in a vertical column followed by different letters are not significantly different (P > 0.05) according to Duncan’s multiple range test
Salicylic acid pretreatment significantly influences morphological changes, chlorophyll, and carotenoid levels in plant leaves under both well-watered and drought-stressed conditions Statistical analysis reveals significant differences at P ≤ 0.05, as determined by Duncan’s multiple range test, with results presented as means ± SE for n = 3.
Drought-stressed plants without salicylic acid (SA) pretreatment exhibited a significant reduction in chlorophyll and carotenoid contents, decreasing by 22.7% and 21.8%, respectively, compared to control plants Conversely, SA-pretreated plants experienced a lesser decrease of only 9.2% in these pigments compared to the control group.
SA pretreatment suppressed oxidative stress induced by drought stress
The O2- content in plants treated with salicylic acid (SA) increased by 56.7% under drought stress, while in drought-stressed plants without SA pretreatment, the O2- content surged by 170% compared to the control group.
28 consistent with accumulation of O2- in situ, indicated by deep brown spots (Figure
Drought stress significantly elevated H2O2 levels, showing a 102.8% increase in non-SA pretreated plants and a 74% increase in SA-pretreated plants compared to control groups Additionally, malondialdehyde (MDA), a byproduct of lipid peroxidation, rose by 52.9% in drought-stressed plants without SA pretreatment, while a 31.6% increase was observed in SA-pretreated plants under similar drought conditions.
Figure 1.2 illustrates the impact of salicylic acid pretreatment on the concentrations of O2- (A), H2O2 (B), and MDA (C), as well as the localization of O2- (D) in the leaves of both control and salicylic acid pretreated plants under well-watered and drought-stressed conditions Significant differences were observed at P ≤ 0.05, as determined by Duncan’s multiple range test, with values expressed as means ± SE for n = 3 The localization of O2- is highlighted by the black areas in the images, which were obtained through histochemical staining using the NBT method and captured at 40X magnification.
In drought-stressed plants, the activities of antioxidant enzymes such as SOD, CAT, GPOD, and APOD increased significantly by 160.9%, 71.8%, 90.6%, and 62.6%, respectively, compared to control plants SA pretreatment further enhanced these enzyme activities, with the exception of GPOD, demonstrating its effectiveness in reducing oxidative stress under drought conditions.
Table 1.2 Changes in the antioxidative system, including superoxide dismutase
This study investigates the activities of superoxide dismutase (SOD), catalase (CAT), guaiacol peroxidase (GPOD), and ascorbate peroxidase (APOD) in the leaves of Chinese cabbage The research focuses on comparing the enzyme activities in plants that were either treated with salicylic acid or left as controls, under both well-watered and drought-stressed conditions.
U: unit Values are presented as means ± SE for n = 3 Values in a vertical column followed by different letters are not significantly different (P > 0.05) according to Duncan’s multiple range test n.s: Non-significant The asterisk indicates significant difference compared with the control: *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001
SA pretreatment is involved in the regulation of redox balance under drought stress conditions
Drought stress led to a significant reduction in reduced glutathione (GSH) levels and a slight increase in oxidized glutathione (GSSG) levels compared to the control However, salicylic acid (SA) pretreatment in drought-stressed plants resulted in a substantial increase in both GSH and GSSG levels, by 65.3% and 70%, respectively The GSH to GSSG ratio decreased by 69.7% in drought-stressed plants without SA pretreatment, but was restored to control levels with SA treatment Additionally, NADPH content decreased by 35% in drought-stressed plants, while it increased by 24.5% and 34.5% in SA-pretreated plants under well-watered and drought conditions, respectively NADP+ levels rose by over 40% in all treatments compared to the control, and the NADPH to NADP+ ratio dropped by 47.3% in drought-stressed plants, with a lesser decrease observed in those treated with SA.
30 pretreated plants with or without drought stress, 31.2% and 24.7%, respectively (Figure 1.3F)
Figure 1.3 Effect of salicylic acid pretreatment on GSH (A), GSSG (B), Ratio of
The study examines the levels of GSH/GSSG (C), NADPH (D), NADP+ (E), and the NADPH/NADP+ ratio (F) in the leaves of control and salicylic acid pretreated plants under both well-watered and drought-stressed conditions Significant differences were observed at P ≤ 0.05, as determined by Duncan’s multiple range test, with results presented as means ± SE for n = 3.
Proline biosynthesis and degradation was regulated by SA pretreatment
Drought stress increased proline content by 231.6% compared to the control Salicylic acid (SA) pretreatment raised proline levels by 96.4% in non-drought-stressed plants and 284.2% in drought-stressed plants, correlating with enhanced expression of proline biosynthesis genes, pyrroline 5-carboxylate synthetase (P5CSA and P5CSB) Specifically, SA pretreatment significantly boosted P5CSA and P5CSB expression by 12.1-fold and 3.0-fold, respectively, compared to controls In contrast, drought stress down-regulated the proline degradation gene, proline dehydrogenase (PDH), with further decreases observed in SA-pretreated plants.
Figure 1.4 illustrates the impact of salicylic acid pretreatment on proline levels and the relative expression of P5CSA, P5CSB, and PDH in the leaves of both control and salicylic acid-treated plants under well-watered and drought-stressed conditions Significant differences were observed at P ≤ 0.05, determined by Duncan’s multiple range test, with results expressed as means ± SE for n = 3.
Introduction
A decrease in water availability significantly affects plant growth by altering metabolite concentrations and disrupting amino acid and carbohydrate metabolism, leading to drought stress This stress response is characterized by the accumulation of reactive oxygen species (ROS) and proline, regulated through hormonal signaling pathways Excessive ROS generation in stressed plants can cause oxidative damage, but they also function as signaling molecules that enhance stress tolerance Proline accumulation serves as a protective mechanism, aiding in the regeneration of NADP+ in chloroplasts, thus mitigating ROS production Studies show that exogenous hydrogen peroxide (H2O2) increases proline levels by activating the enzyme pyrroline-5-carboxylate synthetase (P5CS), while proline treatment can induce calcium-dependent ROS accumulation Phytohormones, particularly abscisic acid (ABA), play a vital role in regulating both ROS generation and proline synthesis, with elevated H2O2 levels promoting ABA-induced proline accumulation, highlighting the intricate interplay between ROS and proline in plant stress responses.
Proline synthesis is triggered by various factors and plays a crucial role in regulating the salt overly sensitive pathway (Chung et al., 2008) Notably, Savouré et al (1997) found that the expression of proline biosynthetic genes is ABA-independent during cold and osmotic stress Additionally, salicylic acid (SA) is significant in managing drought responses, leading to recent studies that explore the complex hormonal interactions through signaling pathways in mutant and transgenic plants (Yasuda et al., 2008; Muñoz-Espinoza et al., 2015) Evidence indicates that an antagonistic relationship between SA and ABA influences plant stress responses (de Torres Zabala et al., 2009; Muñoz-Espinoza et al., 2015).
Salicylic acid (SA) acts as a signal for various stress types, with studies indicating that reactive oxygen species (ROS) mediate SA biosynthesis via calcium signaling and proline-mediated NDR1-dependent pathways Despite these findings, the role of SA in proline synthesis and its importance in cellular redox control, especially regarding drought resistance, remains inadequately understood.
This study explores the effects of salicylic acid (SA) pretreatment and drought conditions on endogenous hormonal status, redox state, reactive oxygen species (ROS), and proline accumulation We hypothesize that these factors respond differently to SA and drought, and that SA-mediated transcriptional regulation, potentially interacting with other hormones, plays a crucial role in enhancing drought tolerance through redox modulation Our investigation focuses on how ROS and proline accumulation, antioxidant activity, and redox status correlate with endogenous hormonal levels and their signaling genes, particularly in the context of redox control mechanisms.
Experiment design
After 6 weeks, plants were selected by morphological similarity and divided into two groups for the salicylic acid (SA) application For SA pretreatment, one group was sprayed daily with 30 mL of 0.5 mM SA based on a preliminary test referring to
In a study conducted by Mateo et al (2006), plants were treated with salicylic acid (SA) and subsequently divided into four treatment groups after five days: non-SA pretreated and well-watered (Control), SA-pretreated and well-watered (SA), non-SA pretreated and drought-stressed (Drought), and SA-pretreated and drought-stressed (SA + Drought) Each group received different irrigation volumes, with well-watered plants receiving 200 mL and drought-stressed plants receiving 20 mL Sampling occurred at five days post-treatment (d 5) and continued at ten days (d 10).
10 days (d 15) after drought treatment (Figure 2)
In this experiment, the response of ROS, antioxidant, proline and redox state response were linked to change in endogenous hormone and their signaling
Figure 2 Experimental design of salicylic acid treated to Brassica napus plants under non-drought and drought stress conditions.
Results
Plants physiological symptom and osmotic potential
Drought stress led to severe stress symptoms in plants, characterized by leaf wilting and a significant decrease in osmotic potential (OP) However, the presence of salicylic acid (SA) during drought conditions mitigated these symptoms, resulting in a lesser decline in OP Additionally, plants pretreated with SA under well-watered conditions showed no changes in their status or OP compared to the control group.
Salicylic acid (SA) pretreatment significantly influences the morphology of Brassica napus plants and their osmotic potential in leaves, particularly under varying water conditions The data, represented as means ± SE for n = 3, indicate that differences marked by distinct letters are statistically significant at P < 0.05, based on Duncan’s multiple range test.
Hormonal status and phytohormone-signaling related gene expression
SA pretreatment significantly increased endogenous SA levels by over three times on day 10, with a slight decrease observed by day 15 under both well-watered and drought-stressed conditions In contrast, drought alone resulted in a notable two-fold increase in SA levels, which was significant only on day 10 Additionally, drought conditions consistently elevated ABA levels, irrespective of SA pretreatment, although the increase in ABA was significantly lower when SA was pretreated.
SA-pretreated plants showed better resilience to drought stress compared to those exposed to drought alone While SA pretreatment resulted in slight changes or no significant increase in JA content, drought stress significantly elevated JA levels compared to the control The ratios of ABA/SA, JA/SA, and (ABA+JA)/SA increased remarkably by 12.1-, 1.6-, and 9.6-fold, respectively, under drought conditions at day 15 compared to the control However, SA pretreatment effectively reduced these ratios to control levels or lower under drought stress.
SA pretreatment significantly enhanced the expression of SA-signaling regulatory genes, including NPR1 and PR1, in both well-watered and drought-stressed plants In contrast, these genes were downregulated in plants subjected to drought conditions alone Additionally, the expression of the ABA synthesis-related gene, NCED3, was also observed.
ABA signaling-related gene, transcriptional factor MYC2, were enhanced by drought alone, whereas their expression levels were alleviated by SA pretreatment
(Figure 2.2C, D) The relative expression of a JA-responsive gene, plant defensin 1.2
(PDF1.2), was up-regulated (+68.1%) by drought stress, but the gene expression was significantly down-regulated (-25.6%) by SA pretreatment at d 15 (Figure 2.2E)
Figure 2.2 illustrates the impact of salicylic acid (SA) pretreatment on the expression of key genes in Brassica napus leaves, including the SA-regulated genes NPR1 and PR1, as well as the ABA synthesis-related gene NCED3, the ABA-signaling gene MYC2, and the JA-signaling gene PDF1.2, under both well-watered and drought stress conditions The data, presented as means ± SE for n = 3, indicate significant differences at P < 0.05, as determined by Duncan’s multiple range test, with bars labeled with different letters reflecting these variations.
Table 2.1 Effects of salicylic acid (SA) pretreatment on hormonal status in leaves of Brassica napus under well-watered or drought-stressed conditions
SA ABA JA SA ABA JA SA ABA JA SA ABA JA
/SA ABA/SA JA/SA (ABA+JA)
Hormone concentrations are expressed in ng g⁻¹ FW, with SA and ABA detailed in Chapter 2 The values presented are means ± SE for n = 3 Significant differences in data are indicated by different letters in the same vertical column at P < 0.05, as determined by Duncan’s multiple range test.
SA reduced oxidative stress under drought stress condition
SA pretreatment resulted in a 2.6-fold increase in O2•− concentration under both well-watered and drought-stressed conditions, with a notable 9.0-fold increase observed in drought conditions alone at day 15 compared to the control This finding was supported by in situ localization of O2•−, revealing a significantly dark spotted area under drought stress Additionally, SA pretreatment alleviated the symptoms caused by drought stress, while H2O2 content showed a significant increase due to either SA pretreatment or drought stress.
Drought stress alone led to the production of H2O2 in plants, while the expression of NAPDH oxidase was significantly increased by over three times with salicylic acid (SA) pretreatment Notably, the drought-induced rise in NAPDH oxidase expression (6.1-fold) was further amplified to 8.8-fold with SA pretreatment by day 15.
Figure 2.3 illustrates the impact of salicylic acid (SA) pretreatment on reactive oxygen species (ROS) accumulation in Brassica napus leaves under both well-watered and drought stress conditions The study highlights the levels of superoxide anion radical (O2 •—) and hydrogen peroxide (H2O2), alongside the expression of the NADPH oxidase gene Notably, O2 •— accumulation is visually represented through nitroblue tetrazolium (NBT) staining under a microscope, with dark areas indicating accumulation (marked by red arrows) The scale bar represents 50mm, and data are reported as means ± SE for n = 3 Significant differences are denoted by different letters at P < 0.05 based on Duncan’s multiple range test.
Superoxide dismutase (SOD) activity was significantly increased by SA pretreatment and/or drought (Figure 2.4A) The higher activity was detected in
44 drought-stressed plant with or without presence of SA Similarly, the expression of
SA pretreatment significantly up-regulated Cu/Zn-SOD and Mn-SOD genes, with higher expression levels observed in SA-pretreated plants compared to those exposed to drought alone Catalase (CAT) activity was notably induced by SA pretreatment on day 10, but it rapidly returned to control levels by day 15 In contrast, CAT activity showed only a slight increase in plants subjected to drought stress alone The expression of CAT was minimally influenced by SA pretreatment and/or drought stress, except for a notable increase in SA-pretreated plants experiencing drought stress on day 10.
Salicylic acid (SA) pretreatment influences the activity of antioxidant enzymes and the expression of their encoding genes in the leaves of Brassica napus under both well-watered and drought stress conditions Notably, the activity of superoxide dismutase (SOD) is affected, along with the expression of SOD-regulated genes such as Cu/Zn-SOD and Mn-SOD Additionally, catalase (CAT) activity is also observed in response to these conditions.
CAT-related gene CAT1 (E) Data are presented as means ± SE for n = 3 Bar labeled with the different letters are significantly different at P < 0.05 according to Duncan’s multiple range test
SA-signaling in regulation redox changes
Salicylic acid (SA) pretreatment significantly increased the levels of NADPH and NADP+ under drought stress conditions While drought alone led to a notable 47.6% decrease in the NADPH/NADP+ ratio, this ratio remained unchanged in SA-pretreated plants compared to the control group Consistent findings were observed in the NADH/NAD+ ratio throughout the experimental period.
Under drought stress, the NADH/NAD+ ratio was reduced by 45% in plants, but this effect was mitigated by salicylic acid (SA) pretreatment Drought stress led to a significant decrease in reduced glutathione (GSH) levels, which returned to control levels following SA pretreatment Additionally, SA-treated plants showed a notable increase in oxidized glutathione (GSSG) content While the GSH/GSSG ratio plummeted by 94.7% in plants subjected to drought alone, it remained stable or only slightly decreased in those pretreated with SA under drought conditions.
Figure 2.5 illustrates the impact of salicylic acid (SA) pretreatment on the expression of redox-regulating genes, including TGA1, WRKY40, TRXh5, and GRXC9, in the leaves of Brassica napus under both well-watered and drought stress conditions The data, represented as means ± SE for n = 3, indicate significant differences at P < 0.05, as determined by Duncan’s multiple range test, with bars labeled with different letters.
Introduction
Drought stress significantly hampers plant growth by limiting water availability, leading to decreased leaf water potential and reduced photosynthetic CO2 assimilation This decline in photosynthetic activity is primarily caused by stomatal closure and chlorophyll degradation Changes in photosynthesis rates and carbon metabolites are crucial for metabolic regulation of stress tolerance and leaf senescence in response to environmental challenges Interestingly, while photosynthesis typically decreases under hydraulic stress, soluble sugar content often increases or remains stable, playing a vital role in osmotic protection The accumulation of soluble sugars helps maintain turgor pressure and hydration during water loss, facilitating osmotic adjustment and enhancing dehydration tolerance Additionally, soluble sugars are implicated in regulating leaf senescence through sugar signaling pathways, while also serving as intermediates in glycolysis to produce carbon skeletons and ATP for anabolic processes.
Sugars, particularly sucrose, play a crucial role in regulating stress responses and tolerance mechanisms in plants The interaction between sugar and hormone signaling is significant in addressing abiotic stress, with specific focus on the relationship between sugar and abscisic acid (ABA) This interaction is believed to trigger various stress responses, such as leaf senescence Additionally, ABA-dependent signaling pathways have been highlighted for their mechanistic role under stress conditions, including ABA-mediated chlorophyll degradation and ABA-responsive element binding (AREB)-mediated sugar accumulation.
In recent studies, ABA-dependent sucrose non-fermenting related kinase (SnRK) activation has been identified as crucial for sucrose metabolism, along with the regulation of leaf starch degradation and the role of ABA-responsive sucrose transporters Additionally, salicylic acid (SA) plays a significant role in both local and systemic resistance against pathogens and helps plants acclimate to abiotic stresses Research indicates that SA is closely associated with photosynthetic performance and related metabolic pathways, highlighting its importance in plant health and stress responses.
SA signaling in the metabolic regulation of sugar accumulation remain to be elucidated in terms of drought tolerance mechanism
This study evaluated the effects of salicylic acid (SA) pretreatment and drought stress on sucrose synthesis, starch degradation, and sugar levels, along with hormonal changes The findings highlight the role of SA in regulating sugar accumulation through transcriptional mechanisms, suggesting a potential interaction with other hormones that enhances drought tolerance.
Experiment design
The experiment, based on the design outlined in Chapter 2, focused on the modification of photosynthetic pigments and sugar status, specifically examining their regulation by endogenous hormones.
Results
SA mitigates drought stress-induced physiological symptoms
Drought stress significantly reduced leaf biomass throughout the experimental period, with a decrease of 45.9% in drought-only conditions compared to a lesser reduction of 22.8% when salicylic acid (SA) was present Leaf water potential was also adversely affected by drought, showing a decline of 66.0% in drought-only plants and 36.0% in SA-pretreated plants at day 15, compared to control The total chlorophyll content experienced the greatest reduction in drought-only conditions Interestingly, SA pretreatment enhanced the expression of the chlorophyll synthase gene (CHLG) despite drought stress, while this gene's expression was slightly decreased in drought-only treatments Additionally, the senescence-associated gene SAG12 was significantly up-regulated under drought stress, showing increases of 4- and 6-fold at days 10 and 15, respectively, with no significant difference in expression observed in SA-pretreated plants under both well-watered and drought-stressed conditions.
The study investigates the impact of exogenous salicylic acid (SA) on the expression of chlorophyll synthase gene (CHLG) and senescence-associated gene 12 (SAG12) in the leaves of both control and SA-pretreated plants under well-watered and drought-stressed conditions Statistical analysis reveals significant differences in gene expression at P ≤ 0.05, as determined by the Duncan multiple range test, with values represented as means ± SE for n = 3.
Salicylic acid (SA) pretreatment significantly influences leaf biomass, water potential, and chlorophyll content in Brassica napus, both under well-watered and drought-stressed conditions The effects of SA on leaf biomass (g per plant, FW), leaf water potential (MPa), and chlorophyll content (mg per g FW) highlight its potential role in enhancing plant resilience to water stress.
Biomass LWP Chlorophyll Biomass LWP Chlorophyll Biomass LWP Chlorophyll Biomass LWP Chlorophyll Control 20.47 -0.55 1.83 36.82 a -0.48 a 1.85 a 62.27 a -0.47 a 1.77 b 64.04 a -0.50 a 1.76 a
LWP: Leaf water potential FW: fresh weight Values are means ± SE for n = 3 Data with different letters in a vertical column are significantly different at P ≤ 0.05 according to Duncan’s multiple range test
SA acts antagonistically to ABA by repressing signaling pathways
Drought stress significantly elevated ABA content in plants, with a notable increase of 12.2-fold in those subjected to drought alone, compared to a 5.3-fold increase in SA-pretreated plants at day 15, relative to the control.
Figure 3.2 illustrates the impact of exogenous salicylic acid (SA) on hormonal status and the expression of signaling-related genes in the leaves of both control and SA-pretreated plants, under well-watered and drought-stressed conditions The analysis includes the content of abscisic acid (ABA) and salicylic acid, alongside the expression levels of key ABA-signaling genes such as sucrose non-fermenting related kinase 2 (SnRK2.2) and ABA-responsive element binding 2 (AREB2), as well as the SA-signaling gene pathogenesis-related gene 2 (PR2) Statistical significance is noted at P ≤ 0.05 based on the Duncan multiple range test, with values presented as means ± SE for n = 3.
Similar to ABA content, the expression of ABA signaling-regulated genes, sucrose non-fermenting related kinase 2 (SnRK2.2) and ABA-responsive element binding 2
Drought significantly enhanced the expression of AREB2 genes in plants, with higher levels observed in those exposed to drought alone compared to SA-pretreated plants Additionally, salicylic acid (SA) pretreatment notably increased endogenous SA levels under both well-watered and drought-stressed conditions throughout the experimental period, while drought alone only elevated SA levels at day 10 These findings align with the observed gene expression patterns.
60 pattern of SA-signaling regulatory genes such as pathogenesis-related (PR) gene 2 (PR2) (Figure 3.2E)
SA induces sugars metabolism and starch degradation
Salicylic acid (SA) pretreatment and drought stress significantly elevated the levels of soluble sugars, including sucrose, glucose, and fructose Sucrose content was found to be 1.6- and 3.2-fold higher in drought conditions and combined SA + drought conditions, respectively, when compared to the control group However, glucose and fructose levels were higher in plants subjected to drought alone than in those pretreated with SA Conversely, starch content decreased notably due to SA pretreatment and/or drought stress, with the lowest levels observed in the SA + drought treatment Throughout the experimental period, SA pretreatment reduced the hexose (glucose + fructose) to sucrose ratio, while drought alone maintained a ratio similar to the control or slightly higher Additionally, the ratio of soluble sugars to starch was significantly increased by SA pretreatment and/or drought stress, with further enhancement in SA-pretreated plants compared to those exposed only to drought.
The study investigates the impact of exogenous salicylic acid (SA) on sucrose phosphate synthase (SPS) activity, cell wall invertase (CWINV) activity, and the expression of the hexokinase 1-related gene (HXK1) in the leaves of both control and SA-pretreated plants under well-watered and drought-stressed conditions Significant differences in the results were observed at P ≤ 0.05, as determined by Duncan's multiple range test, with values presented as means ± SE for n = 3.
The study investigates the impact of exogenous salicylic acid (SA) on the expression of starch degradation enzyme-related genes, specifically β-amylase 1 (BAM1) and α-amylase 3 (AMY3), in the leaves of both control and SA-pretreated plants under varying water conditions Significant differences were observed at P ≤ 0.05, as determined by the Duncan multiple range test, with results expressed as means ± SE for n = 3.
The activity of sucrose phosphate synthase (SPS) significantly increased with salicylic acid (SA) pretreatment and drought, peaking in SA-pretreated plants under drought conditions A similar increase was noted in cell wall invertase activity (CWINV) Conversely, the expression of the hexokinase 1 gene (HXK1), which acts as a sugar sensor, was notably reduced by SA pretreatment and drought, with lower levels in drought-only exposed plants compared to those pretreated with SA Additionally, the expression of starch degradation-related genes, β-amylase 1 (BAM1) and α-amylase 3 (AMY3), was significantly elevated by SA pretreatment in both well-watered and drought-stressed conditions, although drought alone also increased their expression, it remained lower than that observed in the SA + Drought scenario.
Table 3.2 Effects of salicylic acid (SA) pretreatment on soluble sugars (mg g -1 FW) and starch (mg g -1 FW) in the leaves of Brassica napus under well-watered or drought-stressed condition
Sucrose Glucose Fructose Sucrose Glucose Fructose Sucrose Glucose Fructose Sucrose Glucose Fructose Control 14.15 31.76 21.76 15.03 b 26.81 a 21.28 a 10.48 c 38.04 b 23.03 a 17.28 d 46.09 d 18.06 c
Fresh weight (FW) was measured, with hexose represented as the sum of glucose and fructose The values presented are means ± SE based on a sample size of n = 3 Statistically significant differences in data are indicated by different letters within the same vertical column, adhering to a significance level of P ≤ 0.05 as determined by Duncan’s multiple range test.
SA enhances sucrose transportation in xylem and phloem
SA pretreatment significantly increased sucrose levels in the xylem of drought-stressed plants, with higher concentrations observed in both treated and untreated groups In the phloem, sucrose content rose fourfold in SA-pretreated plants under both well-watered and drought conditions at day 15, compared to control plants, while drought-only plants showed a 1.8-fold increase Additionally, SA pretreatment boosted the expression of sucrose transporter genes SUT1, SUT2, and SUT4 throughout the experimental period, with peak expression levels at day 10 In contrast, drought-only plants exhibited a decrease in gene expression by day 15, except for SUT2, which maintained elevated levels.
Figure 3.5 illustrates the impact of exogenous salicylic acid (SA) on sucrose transportation in the leaves of both control and SA-pretreated plants, analyzed under well-watered and drought-stressed conditions The results show sucrose content variations in the xylem (A) and phloem (B), alongside the relative gene expression levels of sucrose transporters SUT1 (C), SUT2 (D), and SUT4 (E) Statistical significance is noted at P ≤ 0.05 based on the Duncan multiple range test, with values presented as means ± SE for n = 3.
SA promotes the contribution of sucrose to osmotic potential
Osmotic potential (OP) significantly decreased by drought stress regardless of SA pretreatment The n of OP was considerably higher in SA-pretreated plants than in
In a study examining the effects of salicylic acid (SA) pretreatment on osmotic potential (OP) in plants, it was found that OP remained unchanged under well-watered conditions compared to the control group However, SA pretreatment significantly enhanced the contribution of sucrose to osmotic potential in both well-watered and drought-stressed plants, unlike the control group Notably, no significant differences were observed in plants subjected solely to drought conditions.
The study investigates the impact of exogenous salicylic acid (SA) on osmotic potential in plant leaves, comparing control plants to those pretreated with SA under both well-watered and drought-stressed conditions Significant differences were observed at P ≤ 0.05, as determined by the Duncan multiple range test, with results presented as means ± SE for n = 3.
Heatmap visualization and Pearson correlation analysis for the metabolites or gene expression
Introduction
Nitrate (NO3-) serves as the primary nitrogen source for plant growth, rapidly stimulating transport and assimilation processes that enhance plant adaptability to changing environments It acts as a nutrient signaling trigger for both local and systemic pathways that regulate carbon and nitrogen metabolism Nitrate assimilation involves the reduction of nitrate to nitrite and ammonium, and it is known to mitigate NH4+ toxicity The regulation of NO3- uptake is linked to the nitrate transceptor gene NRT1.1, which interacts with complex Ca2+-sensor protein kinases and NIN-LIKE PROTEIN (NLP) transcription factors, influencing hormonal responses Additionally, nitrate plays a crucial role in long-distance signaling mechanisms, with nitrate-responsive genes being regulated by hormonal and carbon signaling pathways Recent findings highlight the interconnection between nitrate uptake and hormone status, emphasizing the importance of hormonal regulation of NO3- transport at the transcriptional level, particularly in relation to cytokinin and auxin networks However, the hormonal regulatory mechanisms governing NO3- acquisition under environmental stress remain to be fully elucidated.
Ammonium (NH4+) serves as a crucial metabolic intermediate in nitrogen sources that enhances plant growth The primary pathway for NH4+ assimilation involves the GS/GOGAT cycle, alongside the conversion of α-ketoglutarate facilitated by glutamate dehydrogenase (GDH), establishing a key metabolic node in the synthesis of glutamate.
Research indicates that cytosolic glutamine synthetase (GS) plays a crucial role in proline synthesis by maintaining a consistent glutamate pool during stress conditions Studies have shown that GS and glutamate synthase (GOGAT) activities increase in response to salt and water stress However, an excess of ammonium (NH4+) can lead to leaf chlorosis.
High levels of NH4+ in hyperammonia can be toxic to plants, yet they also promote NH4+ assimilation for glutamate production, serving as a detoxification mechanism (Skopelitis et al., 2006; Jian et al., 2018) In roots, NH4+ boosts drought-induced ABA accumulation, enhancing aquaporin-mediated water uptake and improving drought tolerance (Liu and von Wirén, 2017) Furthermore, glutamate derived from NH4+ assimilation acts as a signaling molecule and a precursor for proline, which is essential for plant development and response to environmental stress (Forde and Lea, 2007; Kan et al., 2017) Adequate glutamate levels are necessary for significant proline accumulation during water stress (Szabados and Savouré, 2010) Research indicates that both glutamate and proline accumulate rapidly in leaves under water-deficit conditions, alongside increased NO3-/NH4+ levels in roots (Chen et al., 2012; Lee et al., 2009b; Zhang et al., 2014; Han et al., 2016) This raises intriguing questions about the distinct roles of NH4+ signaling in relation to ROS and stress hormonal signals during drought stress, which remain largely speculative.
Therefore, a plausible hypothesis given in this work is that hormones couples
The signaling of H2O2 and Ca2+ plays a crucial role in regulating nitrogen transport and assimilation, which in turn facilitates proline accumulation during drought stress This study investigates the relationship between reactive oxygen species (ROS), proline accumulation, nitrogen assimilation enzyme activity, and nitrogen status in response to salicylic acid (SA) and drought conditions, with or without the application of exogenous H2O2 The findings suggest a connection between these responses and the levels of endogenous hormones and their associated signaling genes.
Experiment design
In the study of plants at the eight-leaves stage, conditions were categorized into well-watered and drought stress environments The well-watered group included a control subgroup that received daily irrigation of 200 mL of water, while another subgroup was treated with 0.1 mM salicylic acid (SA), 1 mM hydrogen peroxide (H2O2), and a combination of both at the same volume Drought stress conditions were established for plants that were typically irrigated.
20 mL for plants with or without 1 mM H2O2
Figure 3 Experimental design of salicylic acid, hydrogen peroxide, and drought stress treated to Brassica napus plants.
Results
Plants physiology and stress symptom development
Plant physiology after 5 d of exposure to exogenous SA, H2O2 showed that no difference (Figure 4.1A-D)
The study investigates the effects of salicylic acid and drought on the morphology, osmotic potential, and hydrogen peroxide levels in the leaves of Brassica napus Results are illustrated through various figures, showing significant changes in plant morphology (A-F), osmotic potential (H), and hydrogen peroxide visualization (I), alongside measured H2O2 content (J) Data are presented as mean ± SE (n = 3), with significant differences indicated by different letters on the columns at P < 0.05, based on Duncan’s multiple range test.
Drought stress significantly intensified stress symptoms in plants, evidenced by leaf wilting and a reduction in osmotic potential (OP) The application of salicylic acid (SA) or hydrogen peroxide (H2O2) treatments led to increased H2O2 accumulation, with levels 20% higher in drought conditions alone compared to the combination of H2O2 and drought These findings indicate that H2O2 treatment alleviated some drought stress symptoms, highlighting its potential role in mitigating the effects of drought on plant health.
H2O2 indicated by dark-brown spots in leaves detected (Figure 4.1I)
Modified hormonal status under different stress conditions
Plants treated with exogenous salicylic acid (SA) or hydrogen peroxide (H2O2) showed increased accumulation of SA, trans-Zeatin, and indole-3-acetic acid (IAA), while abscisic acid (ABA) levels were only elevated in SA-treated plants In the presence of SA and H2O2, the levels of these hormones were either unchanged or showed less increase compared to the control group Drought stress significantly raised ABA levels by 13.1-fold, while the increases in SA and trans-Zeatin were lower than those of ABA Furthermore, hormone levels were reduced in the H2O2 + drought condition compared to drought alone, with no significant difference in IAA content observed under drought conditions, regardless of H2O2 treatment.
H2O2 compared to the control (Figure 4.2) The pattern of the SA- and ABA-synthesis related gene expression similar to SA and ABA levels (Figure 4.2E, F)
The study examines the hormonal changes in the leaves of B napus when exposed to salicylic acid (SA) or drought conditions, with and without hydrogen peroxide (H2O2) It highlights the effects of various hormones, including abscisic acid (ABA), trans-Zeatine, and indole-3-acetic acid (IAA), alongside the relative gene expression of SA-synthesis gene ICS1 and ABA-synthesis gene NECD3 Results are presented as mean ± SE (n = 3), with significant differences indicated by different letters on the columns at P < 0.05, based on Duncan’s multiple range test.
In the context of hormonal responses to different treatments, oxidative signal-inducible 1 (OXI1) and mitogen-activated protein kinases 6 (MAPK6) play crucial roles as transduction signals in the hormonal defense pathway These genes exhibit increased expression levels when exposed to exogenous salicylic acid (SA) and hydrogen peroxide (H2O2), particularly in drought-stressed plants However, their expression is slightly diminished in conditions where both SA and H2O2 are present alongside drought stress.
The upregulation of sucrose nonfermenting-1-related protein kinase 2 (SnRK2) enhances ABA levels, which are notably countered by ABA-insensitive 2 (ABI1) Additionally, the expression of nonexpressor pathogenesis-related gene 1 (NPR1) in salicylic acid (SA) regulation, along with cytokinin-binding histidine kinase 5 (CHK5) in trans-Zeatine, exhibits a pattern of expression akin to hormonal-dependent responses.
The expression of hormone defense-related genes in the leaves of Brassica napus was analyzed under various conditions, including exposure to salicylic acid and drought, with or without H2O2 Key genes examined included Oxidative signal-inducible 1 (OXI1), mitogen-activated protein kinase 6 (MAPK6), nonexpressor of pathogenesis-related genes (NPR1), sucrose nonfermenting-1-related protein kinase 2 (SnRK2), ABA-insensitive signal 1 (ABI1), and cytokinin in CHASE receptor kinase 5 (CHK5) The results are presented as mean ± SE (n = 3), with significant differences indicated by different letters on the columns at P < 0.05, as determined by Duncan’s multiple range test.
Change in nitrogen status under drought stress condition
Throughout the experiment, nitrate (NO3-) levels were significantly influenced by different treatments, with elevated concentrations observed in plants treated with salicylic acid (SA) and hydrogen peroxide (H2O2) Notably, drought conditions led to a 120% increase in NO3- compared to control levels In contrast, ammonium (NH4+) concentrations were significantly reduced by H2O2 treatment, showing a 94% decrease under drought stress There was no significant difference in NH4+ levels with or without SA treatment when compared to the control These varying responses of nitrate and ammonium in leaves highlight the role of enzymes in the nitrogen assimilation pathway.
In nitrate assimilation, low nitrate reductase (NR) activity observed in SA- and
In H2O2-treated plants, drought stress resulted in a significant 45% reduction in NR activity compared to the control group Notably, the combination of SA and H2O2 did not influence NR activity When examining ammonium assimilation, the activities of the enzymes GS, GOGAT, and GDH, which are responsible for producing glutamine (Gln) and glutamate (Glu), were analyzed The activity of GS was markedly decreased under drought stress, regardless of H2O2 presence, while other treatments had no significant effect on this enzyme's activity Additionally, GOGAT activity was observed to be lower than expected.
In comparison to the control group, plants treated with H2O2 and subjected to drought conditions showed significant differences, with the exception of those treated with salicylic acid (SA) Notably, GDH activity was elevated by 148.7% in the SA treatment compared to the control Additionally, GDH activity in drought-exposed plants, whether treated with H2O2 or not, increased significantly, while no changes were observed in the SA + H2O2 treatment.
Figure 4.4 illustrates the nitrate and ammonium levels, along with enzyme activity related to nitrogen assimilation pathways in the leaves of B napus under varying conditions of salicylic acid and drought, with or without H2O2 The study focuses on key assimilation enzymes, including nitrate reductase, glutamate synthetase (GS), glutamine oxoglutarate aminotransferase (GOGAT), and glutamate dehydrogenase (GDH) Results are presented as mean ± SE (n = 3), with significant differences indicated by different letters on columns at P < 0.05, as determined by Duncan’s multiple range test.
Calcium signals and glutamate receptor response
To procedure H2O2-linked NADPH oxidase was up-regulated by exogenous SA and
H2O2 Significant differences in NADPH oxidase expression between SA and H2O2
NR: Nitrate reductase activity (nmol NO 2 - mg -1 protein) GS: Glutamine synthase activity (Unit mg -1 protein)
GOGAT: Glutamate synthase activity (nmol NADH mg -1 protein min -1 )
GDH: Glutamate dehydrogenase activity (nmol NADH mg -1 protein min -1 )
79 treated and control plants occurred at day 5 treatments, which was much for drought stress regardless of H2O2-presence (Figure 4.5A)
The study investigates the oxidative burst, calcium ion (Ca 2+) levels, and glutamate receptor response in the leaves of Brassica napus subjected to salicylic acid or drought conditions, with or without hydrogen peroxide (H2O2) Key components analyzed include NADPH oxidase, cytosolic Ca 2+, calcium-dependent protein kinase 5 (CPK5), and glutamate receptor 1.3 (GLR1.3) Results are presented as mean ± SE (n = 3), with significant differences indicated by different letters on the columns at P < 0.05, as determined by Duncan’s multiple range test.
The cytosolic Ca²⁺ concentration increased by 58.8% following treatments with salicylic acid (SA) or hydrogen peroxide (H₂O₂), but no significant difference was observed in the combination of SA and H₂O₂ compared to the control In contrast, drought alone resulted in a substantial 147% increase in Ca²⁺ levels, which was higher than those recorded in the H₂O₂ + drought treatment Throughout the experimental period, the expression of the calcium signaling-related gene calcium-dependent protein kinase 5 (CPK5) was significantly induced by all treatments, with the highest levels noted under drought stress Additionally, the glutamate receptor GLR1.3 was markedly up-regulated in plants treated with SA and/or H₂O₂, exhibiting a seven-fold increase due to drought stress, regardless of H₂O₂ treatment, when compared to the control.
Nitrate and ammonium transporter genes responsese
In the transportation of NO3- is encoded by nitrate transporter gene NRT1.5 in xylem loading and NRT1.7 in phloem loading These genes were downregulated by
SA, H2O2 treated plants Drought stress largely suppressed NRT1.5 and NRT1.7 expression by -0.9-fold and -0.8-fold compared to the control, respectively Two
80 genes less effected by H2O2 + Drought treatment (Figure 4.6E, F) Similar to NH4+ transporter AMT1.1 was suppression in all of the treatments compared to the control, specific to drought stress (Figure 4.6G)
The study investigates the expression of calcium sensor signaling and transporter-related genes in the leaves of B napus under various conditions, including exposure to salicylic acid and drought, with or without H2O2 Key genes analyzed include calcineurin B-like (CBL), CBL-interacting protein kinases (CIPK11/23), TGA1, nitrate transporters (NRT1.5 and NRT1.7), and ammonium transporter 1.1 (AMT1.1) Results are presented as mean ± SE (n = 3), with significant differences indicated by different letters on columns at P < 0.05, as determined by Duncan’s multiple range test.
Calcineurin B-like interacting protein kinases (CIPKs), specifically CIPK11, CIPK23, and calcineurin B-like binding protein 9 (CBL9), play a crucial role in nitrogen transport The expression levels of CBL9, CIPK11, and CIPK23 were found to be slightly elevated by salicylic acid (SA), both in the presence and absence of hydrogen peroxide (H2O2) Notably, under drought stress conditions, the expression of these three genes significantly increased compared to the control, regardless of H2O2 treatment Additionally, the expression of the transcription factor TGA1-dependent was also noted.
Ca 2+ signal was upregulated by SA and/or H2O2 treatment and upon to drought stress (Figure 4.6D)
Change in proline metabolism response to drought stress
Discussion
Proline is a key compatible solute that helps plants adapt to abiotic stress, particularly under water stress conditions Research has shown that nitrate (NO3-) and ammonium (NH4+) play significant roles in proline accumulation during such stress (Kim et al., 2004; Lee et al., 2009b, 2013, 2015) Additionally, the interaction between proline, reactive oxygen species (ROS), and abscisic acid (ABA) is crucial for regulating water stress responses in plants (Verslues et al., 2007; Liang et al., 2013; La et al., 2019) Therefore, it is important to investigate whether changes in ABA content are related to nitrogen status.
To insight into NH4+ assimilation contributes to enhancing proline synthesis in ABA signaling deserves further characterization The present study assessed
84 preferentially the effect of ABA signaling in SA, H2O2 and drought stress treatment to NO3- and NH4+ transport and assimilation
H 2 O 2 is in part of hormone accumulation response to stress
This study emphasizes the role of stress hormones, specifically abscisic acid (ABA) and salicylic acid (SA), in drought tolerance, rather than focusing on plant growth hormones like trans-Zeatine and IAA Our findings indicate that hormonal levels of SA, hydrogen peroxide (H2O2), and drought stress were significantly altered Typically, plants respond to stress through a hypersensitive reaction involving the interplay between H2O2 and phytohormone signaling, as noted in previous research (Xia et al., 2015; Choudhury et al., 2017; Hieno et al.).
In 2019, it was discovered that H2O2 stimulates a signaling pathway that enhances hormone production, leading to increased cytosolic Ca2+ levels (Jiang and Zhang, 2003; Seyfferth and Tsuda, 2014; Osakabe et al., 2014) Treatment with SA and H2O2 resulted in NADPH oxidase activation and a corresponding rise in stress hormones like ABA and SA, along with elevated cytosolic Ca2+ and protein kinase CPK5 expression, particularly under drought stress conditions (Boudsocq and Sheen 2013; Rejeb et al 2015; Stael et al 2015) The proposed model suggests that reactive oxygen species (ROS) interact with Ca2+ signaling and protein kinases in hormone defense, involving Ca2+-dependent protein kinases (CPKs) and oxidative signal-inducible 1 (OXI1) (Foyer and Noctor, 2005) This interaction enhances the activity of NADPH oxidase, promoting the generation of apoplastic ROS (Rentel et al., 2004; Boudsocq and Shen 2013; Dubiella et al 2013) Furthermore, the activation of mitogen-activated protein kinases 6 (MAPK6) is crucial for signal transduction in response to H2O2, which leads to the expression of immunity-related genes (Petersen et al., 2009; Stael et al 2015) Consequently, the synthesis of ABA and SA-related genes (NECD3 and ICS1) and inducible OXI1 is required for MAPK6 action in response to H2O2 generation Notably, in H2O2 + drought conditions, only ABA and SA levels increased, while trans-Zeatine and IAA showed no significant changes.
Under well-watered conditions, H2O2 accumulation was lower compared to drought stress conditions, indicating that certain late H2O2 responses necessitate ABA levels akin to those reported by Hieno et al (2019) Additionally, numerous studies have indicated that reactive oxygen species (ROS) signaling activates the ABA response by promoting ABA synthesis and signaling pathways (Verslues et al., 2007; Černý et al.).
In the ABA signaling pathway, the crosstalk with intracellular H2O2 levels precedes the expression of SnRK2.2, leading to a down-regulation of ABA under H2O2 and drought conditions This indicates that phytohormone signaling is partially mediated by H2O2 responsiveness Our findings demonstrate that exogenous salicylic acid (SA) promotes the accumulation of ABA and trans-Zeatine, which correlates with the activation of ABA receptors (SnRK2.2) and trans-Zeatine receptors (CHK5), alongside the involvement of Ca2+/CPK5 pathways.
In this study, it was clarified that salicylic acid (SA) promotes the expression of NADPH oxidase in a manner dependent on hydrogen peroxide (H2O2) production This process may induce the production of abscisic acid (ABA) and enhance the expression of the ABA-responsive gene SnRK2.2 Additionally, the expression level of NPR1, which acts as the master switch for the SA response, increases alongside elevated endogenous H2O2 levels Conversely, lower levels of ABA and its signaling component SnRK2.2 were observed in treatments combining SA and H2O2 This interplay creates a positive feedback loop that contributes to the sustained relationship between H2O2 and the signaling pathways involved.
Salicylic acid (SA) can enhance the abscisic acid (ABA) response by stimulating SA-responsive pathways through NPR1 Additionally, the hydrogen peroxide (H2O2) signal integrates upstream of SA, facilitating crosstalk via NPR1 signaling This research highlights the role of oxidative stress in these signaling interactions (Herrera-Vásquez et al., 2015; La et al., 2019).
Hydrogen peroxide (H2O2) plays a crucial role in regulating hormone responses in plants under stress, particularly by coupling signals between salicylic acid (SA), abscisic acid (ABA), and H2O2 This indicates that the simultaneous accumulation of these compounds may be significant for plant stress responses.
Hydrogen peroxide (H2O2) and salicylic acid (SA) play a crucial role in the production of abscisic acid (ABA), which is vital for cross-tolerance and signaling pathways Research by Wang et al (2018) indicates that the accumulation of intracellular ABA significantly mitigates SA-induced freezing tolerance This study also confirms that treatment with SA and H2O2 leads to a downregulation of ABA signaling.
The expression of SnRK2.2, as highlighted in our previous study (La et al., 2019), underscores its significance in the response to various stressors This raises an intriguing question about how the specificity of the response is maintained when utilizing a common signaling molecule such as H2O2.
The findings by Mittler et al (2011) highlight the essential role of hydrogen peroxide (H2O2) in facilitating communication between phytohormones that respond rapidly to stress and in reducing stress symptoms, such as alleviating H2O2 effects and delaying leaf senescence (Figure 4.1E, F, I) However, a deeper exploration into the stress-signaling networks and hormonal responses involving H2O2 in stress resistance is necessary for a comprehensive understanding.
Nitrate plays a crucial role in nitrogen assimilation and is involved in signaling transduction, significantly influencing downstream physiological responses in plants through hormones Abscisic acid (ABA), known as a stress hormone, is linked to nitrate signaling, highlighting the connection between ABA accumulation and nitrate response Research indicates that the crosstalk between nitrate and hormonal signals regulates gene expression, demonstrating that plant responses are not solely dependent on nitrate but also on hormonal interactions This is facilitated by nitrate transport and assimilation mechanisms, where nitrate reductase (NR) catalyzes the reduction of nitrate to nitrite, a precursor for ammonium Additionally, nitrate status is managed by nitrate transporter genes (NRT), with NRT1.5 and NRT1.7 identified as key players in long-distance nitrate transport.
Nitrate (NO3-) availability plays a crucial role in plant growth and stress tolerance, as demonstrated by various studies (Chen et al., 2012; Zhang et al., 2014; Han et al., 2016) Research by Chen et al (2012) highlighted that different nrt1.5 mutants indicate that elevated NO3- levels in roots significantly influence plant development Furthermore, the current study reveals that NO3- levels are stimulated in leaves when treated with salicylic acid (SA).
Hydrogen peroxide (H2O2) treatments, along with increased levels of drought stress, reveal that drought-enhanced abscisic acid (ABA) and salicylic acid (SA) are compatible with nitrate (NO3-) restoration in leaves This is achieved through the repression of nitrate assimilation-dependent nitrate reductase activity (NR) and the expression of xylem-phloem nitrate transport genes NRT1.5 and NRT1.7 These findings suggest that ABA and/or SA play a crucial role in regulating these processes during drought stress.
NO3- availability, however, the networks involved in NO3 hormonal crosstalk is still
Nitrate signaling and its impact on protein phosphorylation, particularly involving the nitrate transporter NPF6.3 and nitrate reductase-responsive genes NIA1/NIA2, are crucial for understanding plant responses to high external nitrate concentrations The interaction between calcium sensors CBL1 and CBL9 with protein kinases CIPK11 and CIPK23 leads to the phosphorylation of these transporters, which ultimately inhibits nitrate transport activity The recruitment of CIPK23 to the plasma membrane by CBL1 facilitates its autophosphorylation and full activation, influencing the transport of ions across the plasma membrane Additionally, studies indicate that the ABA-insensitive 1 (ABI1) protein is modulated by ABA or CPK-NLP networks, playing a significant role in the comprehensive response to nitrate The phosphorylation dynamics within the CIPK-CBL complex also highlight the regulatory effects of ABA on members of the clade-A PP2C family, particularly ABI1 and ABI2.
Papers published in scientific journals
A study by Van Hien La et al (2019) published in Environmental and Experimental Botany highlights the role of hormonal shifts in drought tolerance of Brassica napus The research demonstrates that a transition from abscisic acid to salicylic acid-mediated sucrose accumulation significantly enhances the plant's ability to withstand drought conditions This finding underscores the importance of understanding plant hormonal interactions in developing drought-resistant crops.
In the study by Van Hien La et al (2019), published in Environmental and Experimental Botany, the authors investigate how salicylic acid influences drought stress responses in Brassica napus The research focuses on key factors such as reactive oxygen species, proline accumulation, and the overall redox state of the plant Their findings contribute to a better understanding of the physiological mechanisms that underlie drought resilience in this important crop species.
Van Hien La, Bok-Rye Lee, Qian Zhang, Sang-Hyun Park, Md Tabibul Islam,
Tae-Hwan Kim (2019) Salicylic acid ameliorates stress tolerance by regulating redox status and proline metabolism under drought stress in Brassica rapa
Hyo Lee, Bok-Rye Lee, Md Tabibul Islam, Van Hien La, Sang-Hyun Park,
Tae-Hwan Kim (2020) highlights the significance of cultivar variation in the hormonal and sugar responses of Brassica napus The study reveals that abscisic acid-responsive sucrose phloem loading during the early regenerative stage plays a crucial role in determining seed yield in field-grown canola.
Environmental and Experimental Botany 169: 103917 (published online)
Md Tabibul Islam, Bok-Rye Lee, Sang-Hyun Park, Van Hien La, Woo-Jin Jung,
Dong-Won Bae and Tae-Hwan Kim (2019) investigated the hormonal regulation of phenolic compounds in two Brassica napus cultivars with differing susceptibility to Xanthomonas campestris pv campestris Their study, published in Plant Science, highlights the accumulation of soluble and cell-wall bound phenolics, emphasizing the role of hormones in plant defense mechanisms against bacterial pathogens The findings contribute to understanding how these compounds can influence plant resilience and disease resistance in agricultural settings.
In their 2019 study, Md Tabibul Islam and colleagues investigated the effects of p-coumaric acid on Brassica napus, revealing that it promotes jasmonic acid-mediated phenolic accumulation This biochemical response enhances the plant's resistance to black rot disease, highlighting the potential of p-coumaric acid as a natural defense mechanism in crops.
Md Tabibul Islam, Bok-Rye Lee, Protiva Rani Das, Van Hien La, Ha-il Jung,
In the study by Tae-Hwan Kim (2018), the focus is on the characterization of phenolic metabolites in Chinese cabbage, specifically those induced by p-coumaric acid The research examines the relationship between these metabolites and the plant's resistance to Xanthomonas campestris pv campestris, a pathogen affecting the crop The findings contribute to a deeper understanding of disease resistance mechanisms in plants, highlighting the role of soluble and cell wall-bound phenolic compounds in enhancing defense against bacterial infections.
Md Tabibul Islam, Bok-Rye Lee, Sang-Hyun Park, Van Hien La, Dong-Won Bae,
Tae-Hwan Kim (2017) Cultivar variation in hormonal Balance is a significant determinant of disease susceptibility to Xanthomonas campestris pv campestris in Brassica napus Frontiers in Plant Science 8: 2121
II Papers submitted in peer-reviewed scientific journals
The study by Van Hien La et al investigates the hormonal regulatory pathways involved in drought responses in Brassica napus, focusing on the role of glutamate-mediated proline metabolism The research highlights the comparative mechanisms by which these hormonal pathways influence proline accumulation, a crucial factor for plant resilience under drought conditions Understanding these interactions can provide insights into improving drought tolerance in crops.
(Re-submitting to Plants (Basel))
Md Al Mamun, Md Tabibul Islam, Bok-Rye Lee, Van Hien La, Dong-Won Bae,
Tae-Hwan Kim's research focuses on the genotypic variation in resistance gene-mediated calcium signaling, which plays a crucial role in hormonal signaling related to effector-triggered immunity and disease susceptibility in the black rot-Brassica napus pathosystem This study is significant for understanding plant responses to pathogens and enhancing disease resistance strategies in crops The findings will be resubmitted to the journal Plants (Basel) for further consideration.
Van Hien La, Bok-Rye Lee, Md Tabibul Islam, Dong-Won Bae, Tae-Hwan Kim
Turnover of glutathione-integrated cysteine accumulation involves the modulation of salicylic acid and abscisic acid in the accumulation of hydrogen peroxide under water-stress
Van Hien La, Bok-Rye Lee, Md Tabibul Islam, Md Al Mamun, Dong-Won Bae,
Tae-Hwan Kim Nitrate and ammonium assimilation-integrated proline accumulation is associated with H2O2 interplay hormonal of the drought stress responses in Brassica napus L.
Md Tabibul Islam, Bok-Rye Lee, Van Hien La, Dong-Won Bae, Woo-Jin Jung,
Tae-Hwan Kim Label-free quantitative proteomics analysis in susceptible and resistant Brassica napus cultivars infected with Xanthomonas campestris pv campestris
Md Tabibul Islam, Bok-Rye Lee, Woo-Jin Jung, Van Hien La, Dong-Won Bae,
Tae-Hwan Kim Evidence of salicylic acid-induced phenolic metabolites and antifungal activity in Chinese cabbage (Brassica rapa var pekinensis)
유채의 식물 호르몬에 의한 가뭄 스트레스 반응 및 저항성 조절 기전
동물산업학과 전남대학교 대학원
(지도교수 김태환)
This study explores the synergistic effects of hormonal regulation on drought stress responses and resistance.
To characterize various biological processes where antagonistic effects occur, it is essential to understand the underlying mechanisms at play.
수행되었다 1 장부터 3 장 까지는, 살리실산 (SA) 가뭄 저항성 조절에 대한
역할을 평가하였다 4 장에서는, 가뭄스트레스에 대한 저항성 기전에서
The hormonal mechanisms involved in nitrogen metabolism encompass not only the synthesis of glutamic acid but also the accumulation of proline.
Từ chương 1 đến chương 3, bài viết đã giải thích vai trò của điều chỉnh phiên mã trung gian SA trong bối cảnh hạn hán.
저항성, 특히 redox 조절에 포함된 다른 호르몬들, ROS, 항산화제, 프롤린,
The study identified specific interactions with sugars, revealing that drought stress is associated with reactive oxygen species (ROS).
프롤린 축적을 야기했을 뿐만 아니라 redox 의 산화 상태를 증가시켰다 1.5 mM
SA 전처리는 배추에서 ascorbate peroxidase 의 활성이 증가하여 O2•-, H2O2 및
The removal of geological oxidation significantly alleviates the negative effects of drought stress.
SA tiền xử lý giúp giảm căng thẳng do hạn hán cho cây trồng bằng cách tăng cường nồng độ prolin và glutathione.
축적을 야기하였다 산화환원 반응 조절에서 SA, ROS 및 프롤린 상호작용의
상세한 근본적인 메커니즘은 유채에서 평가되었다.0.5mM SA 처리는 가뭄에
의해 유도된 O2•- 축적을 저감시켰으나, H2O2 에는 영향을 주지 못했다 SA 매개
NPR1 은 TRXh5 와 GRXC9 redox 신호 전사를 조절하였고, 이 신호에 의하여
SA 는 redox 상태를 재설정하고, 프롤린 합성 증가 및 ABA 및/또는 JA 신호의
길항적 감소를 보여주었다 반면에, 가뭄은 주로 hexokinase 유전자인 HXK1
Papers in preparation
Van Hien La, Bok-Rye Lee, Md Tabibul Islam, Dong-Won Bae, Tae-Hwan Kim
Turnover of glutathione-integrated cysteine accumulation involves the modulation of salicylic acid and abscisic acid in the accumulation of hydrogen peroxide under water-stress
Van Hien La, Bok-Rye Lee, Md Tabibul Islam, Md Al Mamun, Dong-Won Bae,
Tae-Hwan Kim Nitrate and ammonium assimilation-integrated proline accumulation is associated with H2O2 interplay hormonal of the drought stress responses in Brassica napus L.
Md Tabibul Islam, Bok-Rye Lee, Van Hien La, Dong-Won Bae, Woo-Jin Jung,
Tae-Hwan Kim Label-free quantitative proteomics analysis in susceptible and resistant Brassica napus cultivars infected with Xanthomonas campestris pv campestris
Md Tabibul Islam, Bok-Rye Lee, Woo-Jin Jung, Van Hien La, Dong-Won Bae,
Tae-Hwan Kim Evidence of salicylic acid-induced phenolic metabolites and antifungal activity in Chinese cabbage (Brassica rapa var pekinensis)
유채의 식물 호르몬에 의한 가뭄 스트레스 반응 및 저항성 조절 기전
동물산업학과 전남대학교 대학원
(지도교수 김태환)
This study explores the synergistic effects of hormonal regulation on drought stress responses and resistance.
To characterize various biological processes where antagonistic effects occur, it is essential to understand the underlying mechanisms involved.
수행되었다 1 장부터 3 장 까지는, 살리실산 (SA) 가뭄 저항성 조절에 대한
역할을 평가하였다 4 장에서는, 가뭄스트레스에 대한 저항성 기전에서
The hormonal regulation of nitrogen metabolism encompasses not only the synthesis of glutamic acid but also the accumulation of proline.
Trong các chương từ 1 đến 3, vai trò của việc điều chỉnh phiên mã trung gian SA đã được giải thích trong bối cảnh của sự hạn hán.
저항성, 특히 redox 조절에 포함된 다른 호르몬들, ROS, 항산화제, 프롤린,
The study identified specific interactions with sugar, revealing that drought stress is associated with the production of reactive oxygen species (ROS).
프롤린 축적을 야기했을 뿐만 아니라 redox 의 산화 상태를 증가시켰다 1.5 mM
SA 전처리는 배추에서 ascorbate peroxidase 의 활성이 증가하여 O2•-, H2O2 및
The removal of geological oxidation significantly reduces the negative impacts of drought stress.
SA tiền xử lý đã làm giảm stress do hạn hán ở cây trồng, giúp tăng cường nồng độ proline và glutathione.
축적을 야기하였다 산화환원 반응 조절에서 SA, ROS 및 프롤린 상호작용의
상세한 근본적인 메커니즘은 유채에서 평가되었다.0.5mM SA 처리는 가뭄에
의해 유도된 O2•- 축적을 저감시켰으나, H2O2 에는 영향을 주지 못했다 SA 매개
NPR1 은 TRXh5 와 GRXC9 redox 신호 전사를 조절하였고, 이 신호에 의하여
SA 는 redox 상태를 재설정하고, 프롤린 합성 증가 및 ABA 및/또는 JA 신호의
길항적 감소를 보여주었다 반면에, 가뭄은 주로 hexokinase 유전자인 HXK1
The reduction in expression has led to an increase in hexose levels, partially due to the high gene expression of ABA-dependent sucrose signaling genes SnRK2.2 and AREB2, which in turn enhances sucrose content.
증가시켰다 SA 전처리한 식물에서 sucrose phosphate synthase (SPS) 의 활성과
전분 분해 효소와 관련된 BAM1 과 AMY3 유전자 발현 증가로 인해 sucrose
축적이 가장 높게 일어났다 이러한 결과들은 sucrose 축적 기전에서 ABA로부터
Sự thay đổi đáng kể trong cơ chế của SA có thể giúp điều chỉnh áp suất thẩm thấu và làm giảm quá trình lão hóa của lá.
것을 보여준다
Chapter 4 integrates the regulation of hormones and nitrogen metabolism through proline accumulation, highlighting its significance in physiological processes.
하였다 가뭄에 의해 유도된 ABA 는 0.1 mM SA 와 0.1 mM H2O2 처리구 보다
ABI1 의 발현을 감소시켰고, 세포질 내 Ca 2+ 의존적인 방식으로 CIPK23 과 CBL9
The complex signals have increased the manifestation of NO3- within the xylem-phloem system.
이동과 NO3- 의 동화작용에 부정적인 영향을 미쳤으며, 부분적으로는 NO3- 의
축적을 증가시켰다 SA, H2O2 및 가뭄 처리는 GDH 활성에 의한 대체
Chu trình GS/GOGAT đã kích hoạt quá trình đồng hóa NH4+, giúp hạn chế sự tích tụ của NH4+ Dưới điều kiện căng thẳng do hạn hán, cơ chế điều chỉnh ABA liên quan đến tín hiệu H2O2 và CBL9 đã được kích hoạt.
관련된 가장 높은 GDH 활성에서 추가적인 프롤린 축적은 글루탐산 합성과
The results indicate that plant hormones are closely linked to the responses of plants under drought stress.
Nghiên cứu cho thấy rằng ABA và nhiều hormone khác có vai trò quan trọng trong việc điều chỉnh quá trình chuyển hóa carbon và nitơ.
The antagonistic interactions of salicylic acid (SA) play a crucial role in enhancing drought resistance in rapeseed by activating reactive oxygen species (ROS), sugars, proline, and redox status.