The transcription factor OsbHLH035 mediates seed germination and enables seedling recovery from salt stress through ABA-dependent and ABA-independent pathways, respectively.
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ABSTRACT: Many transcription factors (TFs), such as those in the basic helix-loop-helix (bHLH) family, are important for regulating plant growth and plant responses to abiotic stress. The expression of OsbHLH035 is induced by drought and salinity. However, its functional role in rice growth, development, and the salt response is still unknown.The bHLH TF OsbHLH035 is a salt-induced gene that is primarily expressed in germinating seeds and seedlings. Stable expression of GFP-fused OsbHLH035 in rice transgenic plants revealed that this protein is predominantly localized to the nucleus. Osbhlh035 mutants show delayed seed germination, particularly under salt-stress conditions. In parallel, abscisic acid (ABA) contents are over-accumulated, and the expression of the ABA biosynthetic genes OsABA2 and OsAAO3 is upregulated; furthermore, compared with that in wild-type (WT) seedlings, the salt-induced expression of OsABA8ox1, an ABA catabolic gene, in germinating Osbhlh035 mutant seeds is downregulated. Moreover, Osbhlh035 mutant seedlings are unable to recover from salt-stress treatment. Consistently, sodium is over-accumulated in aerial tissues but slightly reduced in terrestrial tissues from Osbhlh035 seedlings after salt treatment. Additionally, the expression of the sodium transporters OsHKT1;3 and 1;5 is reduced in Osbhlh035 aerial and terrestrial tissues, respectively. Furthermore, genetic complementation can restore both the delayed seed germination and the impaired recovery of salt-treated Osbhlh035 seedlings to normal growth.OsbHLH035 mediates seed germination and seedling recovery after salt stress relief through the ABA-dependent and ABA-independent activation of OsHKT pathways, respectively.
The transcription factor OsbHLH035 mediates seed germination and enables seedling recovery from salt stress through ABA-dependent and ABA-independent pathways, respectively.
<h4>Background</h4>Many transcription factors (TFs), such as those in the basic helix-loop-helix (bHLH) family, are important for regulating plant growth and plant responses to abiotic stress. The expression of OsbHLH035 is induced by drought and salinity. However, its functional role in rice growth, development, and the salt response is still unknown.<h4>Results</h4>The bHLH TF OsbHLH035 is a salt-induced gene that is primarily expressed in germinating seeds and seedlings. Stable expression of ...[more]
Project description:Plant WRKY transcription factors play crucial roles in plant growth and development, as well as plant responses to biotic and abiotic stresses. In this study, we identified and characterized a WRKY transcription factor in rice, OsWRKY50. OsWRKY50 functions as a transcriptional repressor in the nucleus. The transcription of OsWRKY50 was repressed under salt stress conditions, but activated after abscisic acid (ABA) treatment. OsWRKY50-overexpression (OsWRKY50-OX) plants displayed increased tolerance to salt stress compared to wild type and control plants. The expression of OsLEA3, OsRAB21, OsHKT1;5, and OsP5CS1 in OsWRKY50-OX were much higher than wild type and control plants under salt stress. Furthermore, OsWRKY50-OX displayed hyposensitivity to ABA-regulated seed germination and seedling establishment. The protoplast-based transient expression system and yeast hybrid assay demonstrated that OsWRKY50 directly binds to the promoter of OsNCED5, and thus further inhibits its transcription. Taken together, our results demonstrate that rice transcription repressor OsWRKY50 mediates ABA-dependent seed germination and seedling growth and enhances salt stress tolerance via an ABA-independent pathway.
Project description:Alfalfa (Medicago sativa L.) is an important legume crop for forage, agriculture, and environment in the world. Ascorbic acid (AsA) plays positive roles in plants. However, its effects on germination and salt-tolerance of alfalfa are unknown. The effects of AsA applications on seed germination and seedling salt-tolerance of alfalfa were investigated. The results revealed that 0.1 and 1 mmol L-1 of exogenous AsA increased germination, amylase, and protease, as well as seedling length, fresh weight (FW), dry weight (DW), and endogenous AsA both in the shoots and roots, except that 1 mmol L-1 AsA reduced the activities of α-amylase, β-amylase and protease on day 3. However, 10 and 100 mmol L-1 AsA inhibited these parameters and even caused serious rot. It indicates that 0.1 mmol L-1 AsA has the optimal effects, whereas 100 mmol L-1 AsA has the worst impacts. Another part of the results showed that 0.1 mmol L-1 AsA not only enhanced stem elongation, FW and DW, but also increased chlorophyll and carotenoids both under non-stress and 150 mmol L-1 NaCl stress. Furthermore, 0.1 mmol L-1 AsA mitigated the damages of membrane permeability, malondialdehyde, and excessive reactive oxygen species (ROS) and ions both in the shoots and roots under 150 mmol L-1 NaCl stress. Hence, 0.1 mmol L-1 AsA improves growth and induces salt-tolerance by inhibiting excessive ROS, down-regulating the ion toxicity and up-regulating the antioxidant system. The principal component analysis included two main components both in the shoots and roots, and it explained the results well. In summary, the optimum concentration of 0.1 mmol L-1 AsA can be implemented to improve the seed germination and seedling growth of alfalfa under salt stress.
Project description:Soybean is an important and staple oilseed crop worldwide. Salinity stress has adverse effects on soybean development periods, especially on seed germination and post-germinative growth. Improving seed germination and emergence will have positive effects under salt stress conditions on agricultural production. Here we report that NaCl delays soybean seed germination by negatively regulating gibberellin (GA) while positively mediating abscisic acid (ABA) biogenesis, which leads to a decrease in the GA/ABA ratio. This study suggests that fluridone (FLUN), an ABA biogenesis inhibitor, might be a potential plant growth regulator that can promote soybean seed germination under saline stress. Different soybean cultivars, which possessed distinct genetic backgrounds, showed a similar repressed phenotype during seed germination under exogenous NaCl application. Biochemical analysis revealed that NaCl treatment led to high MDA (malondialdehyde) level during germination and the post-germinative growth stages. Furthermore, catalase, superoxide dismutase, and peroxidase activities also changed after NaCl treatment. Subsequent quantitative Real-Time Polymerase Chain Reaction analysis showed that the transcription levels of ABA and GA biogenesis and signaling genes were altered after NaCl treatment. In line with this, phytohormone measurement also revealed that NaCl considerably down-regulated active GA1, GA3, and GA4 levels, whereas the ABA content was up-regulated; and therefore ratios, such as GA1/ABA, GA3/ABA, and GA4/ABA, are decreased. Consistent with the hormonal quantification, FLUN partially rescued the delayed-germination phenotype caused by NaCl-treatment. Altogether, these results demonstrate that NaCl stress inhibits soybean seed germination by decreasing the GA/ABA ratio, and that FLUN might be a potential plant growth regulator that could promote soybean seed germination under salinity stress.
Project description:BACKGROUND: The plant hormone abscisic acid (ABA) regulates diverse processes of plant growth and development. It has recently been proposed that GCR2 functions as a G-protein-coupled receptor (GPCR) for ABA. However, the structural relationships and functionality of GCR2 have been challenged by several independent studies. A central question in this controversy is whether gcr2 mutants are insensitive to ABA, because gcr2 mutants were shown to display reduced sensitivity to ABA under one experimental condition (e.g. 22 degrees C, continuous white light with 150 micromol m(-2) s(-1)) but were shown to display wild-type sensitivity under another slightly different condition (e.g. 23 degrees C, 14/10 hr photoperiod with 120 micromol m(-2) s(-1)). It has been hypothesized that gcr2 appears only weakly insensitive to ABA because two other GCR2-like genes in Arabidopsis, GCL1 and GCL2, compensate for the loss of function of GCR2. PRINCIPAL FINDINGS: In order to test this hypothesis, we isolated a putative loss-of-function allele of GCL2, and then generated all possible combinations of mutations in each member of the GCR2 gene family. We found that all double mutants, including gcr2 gcl1, gcr2 gcl2, gcl1 gcl2, as well as the gcr2 gcl1 gcl2 triple mutant displayed wild-type sensitivity to ABA in seed germination and early seedling development assays, demonstrating that the GCR2 gene family is not required for ABA responses in these processes. CONCLUSION: These results provide compelling genetic evidence that GCR2 is unlikely to act as a receptor for ABA in the context of either seed germination or early seedling development.
Project description:H(2)O(2) is known as a signal molecule in plant cells, but its role in the regulation of aqbscisic acid (ABA) and gibberellic acid (GA) metabolism and hormonal balance is not yet clear. In this study it was found that H(2)O(2) affected the regulation of ABA catabolism and GA biosynthesis during seed imbibition and thus exerted control over seed dormancy and germination. As seen by quantitative RT-PCR (QRT-PCR), H(2)O(2) up-regulated ABA catabolism genes (e.g. CYP707A genes), resulting in a decreased ABA content during imbibition. This action required the participation of nitric oxide (NO), another signal molecule. At the same time, H(2)O(2) also up-regulated GA biosynthesis, as shown by QRT-PCR. When an ABA catabolism mutant, cyp707a2, and an overexpressing plant, CYP707A2-OE, were tested, ABA content was negatively correlated with GA biosynthesis. Exogenously applied GA was able to over-ride the inhibition of germination at low concentrations of ABA, but had no obvious effect when ABA concentrations were high. It is concluded that H(2)O(2) mediates the up-regulation of ABA catabolism, probably through an NO signal, and also promotes GA biosynthesis. High concentrations of ABA inhibit GA biosynthesis but a balance of these two hormones can jointly control the dormancy and germination of Arabidopsis seeds.
Project description:Fatty acid desaturases play important role in plant responses to abiotic stresses. However, their exact function in plant resistance to salt stress is unknown. In this work, we provide the evidence that FAD2, an endoplasmic reticulum localized ?-6 desaturase, is required for salt tolerance in Arabidopsis. Using vacuolar and plasma membrane vesicles prepared from the leaves of wild-type (Col-0) and the loss-of-function Arabidopsis mutant, fad2, which lacks the functional FAD2, we examined the fatty acid composition and Na+-dependent H+ movements of the isolated vesicles. We observed that, when compared to Col-0, the level of vacuolar and plasma membrane polyunsaturation was lower, and the Na+/H+ exchange activity was reduced in vacuolar and plasma membrane vesicles isolated from fad2 mutant. Consistent with the reduced Na+/H+ exchange activity, fad2 accumulated more Na+ in the cytoplasm of root cells, and was more sensitive to salt stress during seed germination and early seedling growth, as indicated by CoroNa-Green staining, net Na+ efflux and salt tolerance analyses. Our results suggest that FAD2 mediated high-level vacuolar and plasma membrane fatty acid desaturation is essential for the proper function of membrane attached Na+/H+ exchangers, and thereby to maintain a low cytosolic Na+ concentration for salt tolerance during seed germination and early seedling growth in Arabidopsis.
Project description:Most crops are sensitive to salt stress, but their degree of susceptibility varies among species and cultivars. In order to understand the salt stress adaptability of Brassica napus to salt stress, we collected the phenotypic data of 505 B. napus accessions at the germination stage under 150 or 215 mM sodium chloride (NaCl) and at the seedling stage under 215 mM NaCl. Genome-wide association studies (GWAS) of 16 salt tolerance coefficients (STCs) were applied to investigate the genetic basis of salt stress tolerance of B. napus. In this study, we mapped 31 salts stress-related QTLs and identified 177 and 228 candidate genes related to salt stress tolerance were detected at germination and seedling stages, respectively. Overexpression of two candidate genes, BnCKX5 and BnERF3 overexpression, were found to increase the sensitivity to salt and mannitol stresses at the germination stage. This study demonstrated that it is a feasible method to dissect the genetic basis of salt stress tolerance at germination and seedling stages in B. napus by GWAS, which provides valuable loci for improving the salt stress tolerance of B. napus. Moreover, these candidate genes are rich genetic resources for the following exploration of molecular mechanisms in adaptation to salt stress in B. napus.
Project description:Sheepgrass is a perennial native grass species in China, and it can tolerate high levels of salt stress with an aggressive and vigorous rhizome system. Many salt-stress-responsive genes have been identified in sheepgrass. In this study, we report the cloning and characterization of a novel salt-induced gene, LcSAIN3 (Leymus chinensis salt-induced 3), from sheepgrass. Expression analysis confirmed that LcSAIN3 was induced by PEG, ABA, and salt treatments, and the expression of LcSAIN3 was significantly increased in salt-tolerant germplasms under salt treatment. Subcellular localization analysis indicated that the GFP-LcSAIN3 protein was mainly localized in the chloroplasts. The heterologous expression of LcSAIN3 in Arabidopsis increased the seed germination rate of transgenic plants under salt, ABA, and mannitol treatments. The seedling survival rate, plant height, and fresh weight of the transgenic plants were higher than those of WT plants under salt stress. The overexpression of LcSAIN3 caused a relatively high accumulation of free proline, enhanced SOD activity, and led to the upregulation of several stress-responsive genes such as AtRD26, AtRD29B, AtSOS1, and AtP5CS1. These results suggest that LcSAIN3 could be a potential target for molecular breeding to improve plants' salt tolerance.
Project description:Urea is intensively utilized as a nitrogen fertilizer in agriculture, originating either from root uptake or from catabolism of arginine by arginase. Despite its extensive use, the underlying physiological mechanisms of urea, particularly its adverse effects on seed germination and seedling growth under salt stress, remain unclear. In this study, we demonstrate that salt stress induces excessive hydrolysis of arginine-derived urea, leading to an increase in cytoplasmic pH within seed radical cells, which, in turn, triggers salt-induced inhibition of seed germination (SISG) and hampers seedling growth. Our findings challenge the long-held belief that ammonium accumulation and toxicity are the primary causes of SISG, offering a novel perspective on the mechanism underlying these processes. This study provides significant insights into the physiological impact of urea hydrolysis under salt stress, contributing to a better understanding of SISG.
Project description:Plant survival depends on seed germination and progression through post-germinative developmental checkpoints. These processes are controlled by the stress phytohormone abscisic acid (ABA). ABA regulates the basic leucine zipper transcriptional factor ABI5, a central hub of growth repression, while the reactive nitrogen molecule nitric oxide (NO) counteracts ABA during seed germination. However, the molecular mechanisms by which seeds sense more favourable conditions and start germinating have remained elusive. Here we show that ABI5 promotes growth via NO, and that ABI5 accumulation is altered in genetic backgrounds with impaired NO homeostasis. S-nitrosylation of ABI5 at cysteine-153 facilitates its degradation through CULLIN4-based and KEEP ON GOING E3 ligases, and promotes seed germination. Conversely, mutation of ABI5 at cysteine-153 deregulates protein stability and inhibition of seed germination by NO depletion. These findings suggest an inverse molecular link between NO and ABA hormone signalling through distinct posttranslational modifications of ABI5 during early seedling development.