Exogenous application of methyl jasmonate alleviates arsenic toxicity by modulating its uptake and translocation in rice (Oryza sativa L.).

Methyl jasmonate (Me-JA) is a plant growth regulator known for modulating plant responses to various abiotic and biotic stresses. The unavoidable arsenic (As) contamination in rice (Oryza sativa) results in reduced crop yield and greater carcinogenic risk to humans. The present work examines the significance of Me-JA induced molecular signaling and tolerance towards arsenic toxicity in rice. The arsenite (AsIII; 25 µM) stress hampered the overall growth and development of the rice seedling. However, the co-application (25 µM AsIII+0.25 µM Me-JA) resulted in increased biomass, chlorophyll content, enhanced antioxidant enzyme activities as compared to AsIII treated plants. The co-application also demonstrated a marked decrease in malondialdehyde content, electrolyte leakage and accumulation of total AsIII content (root + shoot) as compared to AsIII treated plants. The co-application also modulated the expression of genes involved in downstream JA signaling pathway (OsCOI, OsJAZ3, OsMYC2), AsIII uptake (OsLsi1, OsLsi2, OsNIP1;1, OsNIP3;1), translocation (OsLsi6, and OsINT5) and detoxification (OsNRAMP1, OsPCS2, and OsABCC2) which revealed the probable adaptive response of the rice plant to cope up arsenic stress. Our findings reveal that Me-JA alleviates AsIII toxicity by modulating signaling components involved in As uptake, translocation, and detoxification and JA signaling in rice. This study augments our knowledge for the future use of Me-JA in improving tolerance against AsIII stress.

Role of apoplastic calcium during germination and initial stages of seedling establishment in Vigna radiata seeds.

Calcium (Ca2+) is implicated in the initial phase of seed germination and seedling establishment. It is stored complexed with phytic acid during seed development and released by phytase action during germination. We observed phytase activity 18 h post-imbibition (PI) in Vigna seeds, while radicle protrusion occurred approximately 12 h PI. Cotyledon protein extracts prepared 4, 8, 16 and 24 h PI, subjected to Ca2+ immobilized metal affinity chromatography (Ca2+ IMAC), revealed the presence of Ca2+ binding proteins (CaBPs), while Ca2+-dependent amylase activity peaked 18 h PI, implying Ca2+ presence before its release from Ca-phytate, indicating an alternative source of Ca2+. Vigna cotyledon cell-wall preparations 4 h and 24 h PI, titrated against alkali, revealed high cation-binding capacity, and seeds 4 h PI demonstrated high rates of H+ extrusion. Ca2+-binding capacity as well as cell-wall bound Ca2+, measured in cotyledon cell-wall preparations from unimbibed seeds as well as seeds 24 h PI, using a novel competitive chelation technique, showed a marked decline in Ca2+ binding capacity, as well as cell-wall bound Ca2+. Imbibition in the presence of chelators, Ca2+-channel blockers, and H+-pump inhibitors, interfered with germination and radical extension. Further, EDTA-treated cotyledon protein extracts separated on Ca2+IMAC showed a larger CaBP peak than control cotyledon extracts. Pooled fractions clearly showed Ca2+-induced extrinsic fluorescence with anilino -napthalene sulfonate. The results strongly implicate the apoplast may be a major source of Ca2+ in the initial phase of germination and seedling establishment in Vigna seeds.

Recent advances in arsenic metabolism in plants: current status, challenges and highlighted biotechnological intervention to reduce grain arsenic in rice.

Arsenic (As), classified as a "metalloid" element, is well known for its carcinogenicity and other toxic effects to humans. Arsenic exposure in plants results in the alteration of the physiochemical and biological properties and consequently, loss of crop yield. Being a staple food for half of the world's population, the consumption of As-contaminated rice grain by humans may pose serious health issues and risks for food security. In this study, we have described the principal understanding of the molecular basis of arsenic toxicity and accumulation in plant parts. We described the measures for decreasing As accumulation in rice and understanding the mechanism and transport of As uptake, its transport from root to shoot to rice grain, its metabolism, detoxification, as well as the mechanisms lying behind its accumulation in rice grains. There are various checkpoints, such as the tuning of AsV/Pi specific Pi transporters, arsenate reductase, transporters that are involved in the efflux of As to either the vacuole or outside the cell, xylem loading, loading and unloading to the phloem, and transporters involved in the loading of As to grain, that can be targeted to reduce As accumulation in rice grain. Genes/proteins involved in As detoxification, particularly the glutathione (GSH) biosynthesis pathway, phytochelatin (PC) synthesis, and arsenic methyltransferase, also provide a great pool of pathways that can also be castellated for the low As in rice grains. Paddy rice is also used as fodder for animals, enhancing vacuolar sequestration and using constitutive promoters, which may be of concern for animal health. Therefore, using a root-specific promoter and/or converting inorganic arsenic into volatile organic arsenic might be a better strategy for low As in grain. Furthermore, in this review, the other specific approaches, such as bio-remediation, bio-augmentation practices, and molecular breeding, which have great potential to reduce As uptake from soil to rice grains, have also been highlighted.

Rice (Oryza sativa L.) tau class glutathione S-transferase (OsGSTU30) overexpression in Arabidopsis thaliana modulates a regulatory network leading to heavy metal and drought stress tolerance.

Glutathione S-transferases (GSTs) are well-known enzymes due to their role in detoxification of xenobiotic compounds. However, their biochemical action is still not so clear in imparting tolerance to several abiotic stresses in crop plants. In our previous study, we observed that rice tau class OsGSTU30 plays a significant role in the detoxification of Cr(vi). Interestingly, q-RT PCR analysis also revealed higher expression of OsGSTU30 under drought conditions. In this study, we characterize OsGSTU30 in response to drought as well as heavy metal [Cr(vi)] stresses through overexpression in Arabidopsis thaliana. Biochemical and physiological analyses revealed that OsGSTU30 overexpression lines have improved tolerance against both stresses as compared to wild-type plants. Kinetic analysis and molecular docking confirmed that OsGSTU30 enzyme possesses both GST as well as glutathione peroxidase (GPx) like activity. Differentially expressed stress-responsive genes were also identified by transcriptome analysis, involved in different biological pathways during abiotic stresses. These results suggest the signaling functions of OsGSTU30 apart from its catalytic activity during abiotic stress responses and can be further exploited for improving the stress tolerance in crops.

Genome-wide analysis of rice dehydrin gene family: Its evolutionary conservedness and expression pattern in response to PEG induced dehydration stress.

Abiotic stresses adversely affect cellular homeostasis, impairing overall growth and development of plants. These initial stress signals activate downstream signalling processes, which, subsequently, activate stress-responsive mechanisms to re-establish homeostasis. Dehydrins (DHNs) play an important role in combating dehydration stress. Rice (Oryza sativa L.), which is a paddy crop, is susceptible to drought stress. As drought survival in rice might be viewed as a trait with strong evolutionary selection pressure, we observed DHNs in the light of domestication during the course of evolution. Overall, 65 DHNs were identified by a genome-wide survey of 11 rice species, and 3 DHNs were found to be highly conserved. The correlation of a conserved pattern of DHNs with domestication and diversification of wild to cultivated rice was validated by synonymous substitution rates, indicating that Oryza rufipogon and Oryza sativa ssp. japonica follow an adaptive evolutionary pattern; whereas Oryza nivara and Oryza sativa ssp. indica demonstrate a conserved evolutionary pattern. A comprehensive analysis of tissue-specific expression of DHN genes in japonica and their expression profiles in normal and PEG (poly ethylene glycol)-induced dehydration stress exhibited a spatiotemporal expression pattern. Their interaction network reflects the cross-talk between gene expression and the physiological processes mediating adaptation to dehydration stress. The results obtained strongly indicated the importance of DHNs, as they are conserved during the course of domestication.

Reactive oxygen species mediate axis-cotyledon signaling to induce reserve mobilization during germination and seedling establishment in Vigna radiata.

Seeds represent an excellent opportunity to investigate the role of reactive oxygen species (ROS) in control of metabolism during germination and seedling establishment. Cotyledons, the storage organs in Vigna, do not display growth/cell division while the embryonic axis shows rapid growth and intense metabolic activity. The present study investigates the possibility of ROS generated during respiration in the axis serving as messengers guiding storage reserve mobilization from cotyledons at the pre-greening stage. Seeds were germinated in the presence of hydroxyurea to halt cell division in the S-phase and separately in Edaravone, a potent free radical scavenger. Both treatments caused a decrease in germination percentage, seedling growth and protein mobilization. In the growing axis, both treatments resulted in a decrease in hydrogen peroxide (H2O2), total ROS, MDA and protein carbonyls. The picture in cotyledons was quite different, owing to the physiological dissimilarities between the tissues. The status of redox as evident by GSH/GSSG ratios tended toward oxidizing in axis in comparison to the highly reducing environment found in cotyledons. This is construed as a tendency to maintain redox buffering on the oxidizing side in the axis, to facilitate the passage of ROS message. These results strongly indicate that suppression of cell division or scavenging of ROS adversely affects protein reserve mobilization. It is proposed that apart from H2O2 being a transportable signal, the final message perceived in cotyledons also comprises lipid peroxidation, protein carbonylation and alteration of redox status of the glutathione pool.

Role of H2O2 and cell wall monoamine oxidases in germination of Vigna radiata seeds.

Plant cell wall expresses monoamine oxidases (MAOs) that catalyze oxidation of secreted amines and produce H2O2 in the process. The H2O2, so produced is used by cell wall peroxidases for lignification of cell wall or for plant defense. The natural substrates for these MAOs are elusive, but polyamines and certain catecholamines have been proposed as candidates. Reactive oxygen species are also known to act as signaling molecules controlling plant metabolism. Mungbean (Vigna radiata) has long served as the plant model of choice while studying molecular programs followed during germination and seed development. In this study, we tested the effect of externally added MAO substrates epinephrine and H2O2 on storage protein mobilization in germinating seeds of Vigna radiata. The seeds were imbibed in the presence of 50 microM epinephrine and 10 microM H2O2. These low concentrations of the two compounds were used to exclude direct effects on proteolysis and were arrived at after testing a range of the two and choosing the most effective concentration. These seeds showed 11% and 7% decrease in fresh weight respectively, indicating greater storage mobilization and a corresponding 19% and 46% increase in axis length as compared to untreated seeds. Soluble protein in seeds treated with epinephrine and H2O2 decreased significantly by 34% and 33% as compared to untreated seeds. Electrophoretic analysis of seed proteins revealed a startling and selective depletion of storage proteins in treated seeds. The results indicated a clear involvement of H2O2 in storage protein mobilization in the cotyledons. We propose that H2O2 generated within cell walls of seeds serves as a signaling molecule guiding germination events, including protein reserve mobilization.

A novel Ca2+-activated protease from germinating Vigna radiata seeds and its role in storage protein mobilization.

Calcium (Ca(2+))-dependent/activated proteases make decisive cleavages in proteins, affecting their further degradation/activation. Few such Ca(2+)-dependent proteases have been reported from plants, and none during germination-related events. Seeds are woken up from their quiescent state upon imbibition of water. The subsequent process of germination is strongly influenced by hormones (mainly gibberellins) and light, with both resulting in change in intracellular Ca(2+). We have investigated the effect of Ca(2+) on protease activity in extracts prepared from dry Vigna radiata (L.) Wilczec seeds and cotyledons 4, 24, 48 and 72h post-imbibition. Ca(2+)-activated protease activity is present at a very low level in dry seeds, rises with imbibition and peaks 24h post-imbibition. Subsequently, the activity rapidly declines, even as total protease activity continues to rise. Calcium activation of proteolysis was reversed by ethylene diamine tetraacetic acid (EDTA), ethylene glycol-bis (2-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA), 1,10, phenanthroline, chlorpromazine and by beta-mercaptoethanol in a concentration-dependent manner. Protease activity was also inhibited by para chloro mercuribenzoate (pCMB) and l-trans-epoxysuccinyl-leucylamido(4-guanidino) butane (E 64), while phenyl methyl sulfonyl fluoride (PMSF) and pepstatin did not effect Ca(2+) activation. The protease could be separated from the calmodulin fraction by size-exclusion chromatography, while retaining its ability for Ca(2+) activation, excluding the possibility of activation through calmodulin-based pathways. The presence of a Ca(2+)-activated protease in the cotyledons suggests its role in a predetermined program of germination involving elevation of cytosolic Ca(2+) levels during germination. This protease could be an important enzyme interfacing cytoplasmic signaling events and initiation of storage protein mobilization during seed germination.