Browsing by Author "Gupta, Kapuganti Jagadis"

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Adaptive reprogramming during early seed germination requires temporarily enhanced fermentation-A critical role for alternative oxidase regulation that concerns also microbiota effectiveness
(Frontiers Media S.A., 2021) Bharadwaj, Revuru; Noceda, Carlos; Mohanapriya, Gunasekharan; Kumar, Sarma Rajeev; Thiers, Karine Leitão Lima; Costa, José Hélio; Macedo, Elisete Santos; Kumari, Aprajita; Gupta, Kapuganti Jagadis; Srivastava, Shivani; Adholeya, Alok; Oliveira, Manuela; Velada, Isabel; Sircar, Debabrata; Sathishkumar, Ramalingam; Arnholdt-Schmitt, Birgit
Plants respond to environmental cues via adaptive cell reprogramming that can affect whole plant and ecosystem functionality. Microbiota constitutes part of the inner and outer environment of the plant. This Umwelt underlies steady dynamics, due to complex local and global biotic and abiotic changes. Hence, adaptive plant holobiont responses are crucial for continuous metabolic adjustment at the systems level. Plants require oxygen-dependent respiration for energy-dependent adaptive morphology, such as germination, root and shoot growth, and formation of adventitious, clonal, and reproductive organs, fruits, and seeds. Fermentative paths can help in acclimation and, to our view, the role of alternative oxidase (AOX) in coordinating complex metabolic and physiological adjustments is underestimated. Cellular levels of sucrose are an important sensor of environmental stress. We explored the role of exogenous sucrose and its interplay with AOX during early seed germination. We found that sucrose-dependent initiation of fermentation during the first 12 h after imbibition (HAI) was beneficial to germination. However, parallel upregulated AOX expression was essential to control negative effects by prolonged sucrose treatment. Early downregulated AOX activity until 12 HAI improved germination efficiency in the absence of sucrose but suppressed early germination in its presence. The results also suggest that seeds inoculated with arbuscular mycorrhizal fungi (AMF) can buffer sucrose stress during germination to restore normal respiration more efficiently. Following this approach, we propose a simple method to identify organic seeds and low-cost on-farm perspectives for early identifying disease tolerance, predicting plant holobiont behavior, and improving germination. Furthermore, the research strengthens the view that AOX can serve as a powerful functional marker source for seed hologenomes.
Alternative oxidase (AOX) senses stress levels to coordinate auxin-induced reprogramming from seed germination to somatic embryogenesis- A role relevant for seed vigor prediction and plant robustness
(Frontiers Media S.A., 2019) Mohanapriya, Gunasekaran; Bharadwaj, Revuru; Noceda, Carlos; Costa, José Hélio; Kumar, Sarma Rajeev; Sathishkumar, Ramalingam; Thiers, Karine Leitão Lima; Macedo, Elisete Santos; Silva, Sofia; Annicchiarico, Paolo; Groot, Steven P.C.; Kodde, Jan; Kumari, Aprajita; Gupta, Kapuganti Jagadis; Arnholdt-Schmitt, Birgit
Somatic embryogenesis (SE) is the most striking and prominent example of plant plasticity upon severe stress. Inducing immature carrot seeds perform SE as substitute to germination by auxin treatment can be seen as switch between stress levels associated to morphophysiological plasticity. This experimental system is highly powerful to explore stress response factors that mediate the metabolic switch between cell and tissue identities. Developmental plasticity per se is an emerging trait for in vitro systems and crop improvement. It is supposed to underlie multi-stress tolerance. High plasticity can protect plants throughout life cycles against variable abiotic and biotic conditions. We provide proof of concepts for the existing hypothesis that alternative oxidase (AOX) can be relevant for developmental plasticity and be associated to yield stability. Our perspective on AOX as relevant coordinator of cell reprogramming is supported by real-time polymerase chain reaction (PCR) analyses and gross metabolism data from calorespirometry complemented by SHAM-inhibitor studies on primed, elevated partial pressure of oxygen (EPPO)–stressed, and endophyte-treated seeds. In silico studies on public experimental data from diverse species strengthen generality of our insights. Finally, we highlight ready-to-use concepts for plant selection and optimizing in vivo and in vitro propagation that do not require further details on molecular physiology and metabolism. This is demonstrated by applying our research & technology concepts to pea genotypes with differential yield performance in multilocation fields and chickpea types known for differential robustness in the field. By using these concepts and tools appropriately, also other marker candidates than AOX and complex genomics data can be efficiently validated for prebreeding and seed vigor prediction.
Alternative oxidase is an important player in the regulation of nitric oxide levels under normoxic and hypoxic conditions in plants
(Oxford University Press, 2019) Kumari, Aprajita; Pathak, Pradeep Kumar; Bulle, Mallesham; Igamberdiev, Abir U; Gupta, Kapuganti Jagadis
Plant mitochondria possess two different pathways for electron transport from ubiquinol: the cytochrome pathway and the alternative oxidase (AOX) pathway. The AOX pathway plays an important role in stress tolerance and is induced by various metabolites and signals. Previously, several lines of evidence suggested that the AOX pathway prevents overproduction of superoxide and other reactive oxygen species. More recent evidence suggests that AOX also plays a role in regulation of nitric oxide (NO) production and signalling. The AOX pathway is induced under low phosphate, hypoxia, pathogen infections and elicitor treatments. The induction of AOX under aerobic conditions in response to various stresses can reduce electron transfer through complexes III and IV and thus prevents the leakage of electrons to nitrite and the subsequent accumulation of NO. Excess of NO under various stresses can inhibit complex IV, thus the AOX pathway minimizes nitrite-dependent NO synthesis that would arise from enhanced electron leakage in the cytochrome pathway. By preventing NO generation, AOX can reduce peroxynitrite formation and tyrosine nitration. In contrast to its function under normoxia, AOX has a specific role under hypoxia, where AOX can facilitate nitrite-dependent NO production. This reaction drives the phytoglobin-NO cycle to increase energy efficiency under hypoxia.
Alternative oxidase plays a role in minimizing ROS and RNS produced under salinity stress in Arabidopsis thaliana
(John Wiley & Sons, 2022) Manbir; Singh, Pooja; Kumari, Aprajita; Gupta, Kapuganti Jagadis
Under stress conditions, the overproduction of different reactive oxygen species (ROS) and reactive nitrogen species (RNS) causes imbalance in the redox homeostasis of the cell leading to nitro-oxidative stress in plants. Alternative oxidase (AOX) is a conserving terminal oxidase of the mitochondrial electron transport chain, which can minimize the ROS. Still, the role of AOX in the regulation of RNS during nitro-oxidative stress imposed by salinity stress is not known. Here, we investigated the role of AOX in minimizing ROS and RNS induced by 150 mM NaCl in Arabidopsis using transgenic plants overexpressing (AOX OE) and antisense lines (AOX AS) of AOX. Imposing NaCl treatment leads to a 4-fold enhanced expression of AOX accompanied by enhanced AOX capacity in WT Col-0. Further AOX-OE seedlings displayed enhanced growth compared with the AOX-AS line under stress. Examination of NO levels by DAF-FM fluorescence and chemiluminescence revealed that AOX overexpression leads to reduced levels of NO. The total NR activity was elevated under NaCl, but no significant change was observed in wild-type (WT), AOX OE, and AS lines. The total ROS, superoxide, H2O2 levels, and lipid peroxidation were higher in the AOX-AS line than in WT and AOX-OE lines. The peroxynitrite levels were also higher in the AOX-AS line than in WT and AOX-OE lines; further, the expression of antioxidant genes was elevated in AOX-AS. Taken together, our results suggest that AOX plays an important role in the mitigation of ROS and RNS levels and enhances plant growth, thus providing tolerance against nitro-oxidative stress exerted by NaCl.
Can we secure food and nutrition through crop innovation ?
(John Wiley & Sons, 2025) Pareek, Ashwani; Gupta, Kapuganti Jagadis; Singla-Pareek, Sneh L.; Foyer, Christine H.
Climate change poses an existential challenge to global food and nutritional security by disrupting agricultural systems, altering crop yields, and affecting the availability of essential nutrients. Increasing temperatures, erratic rainfall patterns, and a greater frequency of extreme weather events negatively impact crop productivity, threatening the stability of food supply chains. Additionally, climate-induced stresses such as drought, salinity, and heat not only reduce yield quantity but also affect the nutritional composition of staple crops, potentially exacerbating micronutrient deficiencies. Addressing these challenges necessitates a multi-faceted approach, integrating genetic improvements, sustainable agricultural practices, and the development of climate-resilient crops that can thrive under adverse conditions. This special issue on ‘Food and Nutritional Security' brings together state-of-the-art reviews by experts and cutting-edge studies that highlight the importance of current research in crop science. The reviews and case studies that are contained in this volume provide a concise overview of the field, acknowledging current gaps in knowledge, while examining the potential of genetic innovations, sustainable agronomic practices, and biotechnological advancements in addressing food security challenges.
Chemiluminescence detection of nitric oxide from roots, leaves, and root mitochondria
(Springer, 2016) Wany, Aakanksha; Gupta, Alok Kumar; Kumari, Aprajita; Gupta, Shika; Mishra, Sonal; Jaintu, Ritika; Pathak, Pradeep K.; Gupta, Kapuganti Jagadis
NO is a free radical with short half-life and high reactivity; due to its physiochemical properties it is very difficult to detect the concentrations precisely. Chemiluminescence is one of the robust methods to quantify NO. Detection of NO by this method is based on reaction of nitric oxide with ozone which leads to emission of light and amount of light is proportional to NO. By this method NO can be measured in the range of pico moles to nano moles range. Using direct chemiluminescence method, NO emitted into the gas stream can be detected whereas using indirect chemiluminescence oxidized forms of NO can also be detected. We detected NO using purified nitrate reductase, mitochondria, cell suspensions, and roots; detail measurement method is described here.
Chickpea chitinases responsive to Helicoverpa herbivory and phytohormone signaling: genome-wide identification, field expression profiling, and structure-guided prioritization
(BioMed Central Ltd, 2026) Konda, Aravind Kumar; Annapragada, Harika; G K, Sujayanand; Singh, Pooja; Bhuvanachandra, Bhoopal; Chinnasamy, Hariharan V; Dixit, Girish Prasad; Gupta, Kapuganti Jagadis; Matheshwaran, Saravanan
Background: Chitinases can contribute to plant defence against fungal pathogens and insect herbivores, but their family organization, inducible deployment, and putative ligand-recognition behaviour remain poorly resolved in chickpea. We combined genome-wide identification, field expression profiling under controlled Helicoverpa armigera infestation, hormone treatments, and structure-guided comparison of representative proteins to prioritize defence-associated chickpea chitinases. Results: We identified 28 chickpea chitinase loci (Car_Chits), comprising 22 glycosyl hydrolase family 18 (GH18) genes and 6 GH19 genes. Local duplication, especially tandem duplication within GH18, was the main contributor to family expansion, and interpretable duplicate pairs were retained mainly under purifying selection. Promoter scans indicated broad enrichment of defence- and hormone-associated cis-elements. Field quantitative real-time PCR (qRT-PCR) profiling of 11 candidate genes in field-grown plants subjected to controlled H. armigera infestation and hormone treatments showed treatment-specific temporal regulation. Car_Chit-4 (GH19) was strongly induced by salicylic acid (7.81-fold at 0.5 h; q < 0.05) but transiently repressed shortly after H. armigera feeding (0.15-fold at 0.5 h; q = 0.030). Car_Chit-19 (GH18) was the clearest herbivory-responsive gene, with late induction at 8 h (1.62-fold; q = 0.050) and 48 h (1.85-fold; q = 0.050). Jasmonic acid caused broad early repression across several genes, followed by delayed induction of Car_Chit-4 at 24 h. Seven Car_Chit-(GlcNAc)₄ complexes were modelled, docked, and simulated for 100 ns. GH18 proteins generally showed more favourable predicted MM-PBSA binding energies than GH19 proteins, but the structural metrics were interpreted as relative ligand-recognition indicators rather than direct evidence of anti-herbivore function. Car_Chit-17 had the most favourable predicted binding energy (ΔG_bind = - 18.51 ± 6.75 kcal/mol), whereas Car_Chit-14 and Car_Chit-27 retained the most stable ligand poses and Car_Chit-19 displayed the most stable protein scaffold. Conclusions: Chickpea chitinases show differentiated temporal responses to herbivory and hormone signalling. The study supports a working model in which GH19 Car_Chit-4 marks a rapid salicylic-acid-responsive arm, whereas GH18 Car_Chit-19 marks a delayed herbivory-responsive arm. A tiered prioritization framework separates expression-deployed candidates from structure-guided biochemical candidates, explaining why different genes emerge from qRT-PCR and molecular modelling analyses. The structural analyses provide complementary prioritization of Car_Chit-17, Car_Chit-14, and Car_Chit-27 for biochemical characterization. Together, these results provide a resource for dissecting chitinase-mediated defence in chickpea and for selecting candidates for functional validation.
Co-overexpression of SWEET sucrose transporters modulates sucrose synthesis and defence responses to enhance immunity against bacterial blight in rice
(John Wiley & Sons, 2024) Singh, Jitender; James, Donald; Das, Shubhashis; Patel, Manish Kumar; Sutar, Rashmi Ranjan; Achary, V. Mohan Murali; Goel, Naveen; Gupta, Kapuganti Jagadis; Reddy, Malireddy K.; Jha, Gopaljee; Sonti, Ramesh V.; Foyer, Christine H.; Thakur, Jitendra K.; Tripathy, Baishnab C.
Enhancing carbohydrate export from source to sink tissues is considered to be a realistic approach for improving photosynthetic efficiency and crop yield. The rice sucrose transporters OsSUT1, OsSWEET11a and OsSWEET14 contribute to sucrose phloem loading and seed filling. Crucially, Xanthomonas oryzae pv. oryzae (Xoo) infection in rice enhances the expression of OsSWEET11a and OsSWEET14 genes, and causes leaf blight. Here we show that co‐overexpression of OsSUT1, OsSWEET11a and OsSWEET14 in rice reduced sucrose synthesis and transport leading to lower growth and yield but reduced susceptibility to Xoo relative to controls. The immunity‐related hypersensitive response (HR) was enhanced in the transformed lines as indicated by the increased expression of defence genes, higher salicylic acid content and presence of HR lesions on the leaves. The results suggest that the increased expression of OsSWEET11a and OsSWEET14 in rice is perceived as a pathogen (Xoo) attack that triggers HR and results in constitutive activation of plant defences that are related to the signalling pathways of pathogen starvation. These findings provide a mechanistic basis for the trade‐off between plant growth and immunity because decreased susceptibility against Xoo compromised plant growth and yield.
Comparative investigation on chemical and green synthesized titanium dioxide nanoparticles against chromium (VI) stress eliciting differential physiological, biochemical, and cellular attributes in Helianthus annuus L.
(Elsevier B.V., 2024) Kumar, Dharmendra; Mariyam, Safoora; Gupta, Kapuganti Jagadis; Thiruvengadam, Muthu; Ghodake, Gajanan Sampatrao; Xing, Baoshan; Seth, Chandra Shekhar
Nanotechnology is a new scientific area that promotes unique concepts to comprehend the optimal mechanics of nanoparticles (NPs) in plants under heavy metal stress. The present investigation focuses on effects of synthetic and green synthesized titanium dioxide nanoparticles (TiO2 NPs and gTiO2 NPs) against Cr(VI). Green TiO2 NPs have been produced from plant leaf extract (Ricinus communis L.). Synthesis was confirmed employing an array of optical spectroscopic and electron microscopic techniques. Chromium strongly accelerated H2O2 and MDA productions by 227 % and 266 % at highest chromium concentration (60 mg/kg of soil), respectively, and also caused DNA damage, and decline in photosynthesis. Additionally, anomalies were observed in stomatal cells with gradual increment in chromium concentrations. Conversely, foliar applications of TiO2 NPs and gTiO2 NPs considerably mitigated chromium stress. Sunflower plants treated with modest amounts of green TiO2 NPs had significantly better growth index compared to chemically synthesized ones. Principal component analysis highlighted the variations among photosynthetic attributes, oxidative stress markers, and antioxidant defense systems. Notably, gTiO2 supplementation to the Cr(VI) strained plants minimized PC3 production which is a rare report so far. Conclusively, gTiO2 NPs have been identified to be promising nano-based nutrition resource for farming applications.
Cross-species expression of OsDJ-1C from rice enhances tolerance to salinity and drought stress in tomato
(Elsevier B.V., 2026) Mishra, Manjari; Chatterjee, Yajnaseni; Gupta, Brijesh Kumar; Tomar, Surabhi; Babuta, Priyanka; Gupta, Kapuganti Jagadis; Pareek, Ashwani; Singla-Pareek, Sneh Lata
Abiotic stresses such as salinity and drought induce the accumulation of methylglyoxal (MG), a highly cytotoxic dicarbonyl compound that disrupts cellular metabolism in plants. MG detoxification is primarily mediated by the glutathione-dependent glyoxalase pathway, classically comprising the enzymes glyoxalase I and II. In contrast, glyoxalase III (GLYIII) catalyzes detoxification of MG in a single-step without requiring glutathione. In the present study, we investigated the functional role of OsDJ-1C, a rice GLYIII enzyme, by heterologous overexpression in tomato (Solanum lycopersicum). Transgenic lines exhibited significantly enhanced stress tolerance through a more efficient antioxidant defense mechanism under stress conditions. This improvement was driven by increased GLYIII-mediated detoxification of MG, leading to effective suppression of reactive oxygen species (ROS) accumulation. Reduced ROS levels in the overexpression lines resulted in greater internal oxygen availability and enhanced cellular respiration than wild-type plants. Furthermore, transgenic plants maintained higher pyruvate levels than the wild-type controls, thereby sustaining tricarboxylic acid (TCA) cycle flux and ATP production under stress. Overall, these findings reveal a conserved, cross-species function of OsDJ-1C in enhancing abiotic stress tolerance emphasizing its relevance for improving agricultural sustainability and food security under changing climatic conditions.
Current approaches to measure nitric oxide in plants
(Oxford University Press, 2019) Vishwakarma, Abhaypratap; Wany, Aakanksha; Pandey, Sonika; Bulle, Mallesham; Kumari, Aprajita; Kishorekumar, Reddy; Igamberdiev, Abir U.; Mur, Luis A.J.; Gupta, Kapuganti Jagadis
Nitric oxide (NO), is now established as an important signalling molecule in plants where they influence growth, development and responses to stress. Despite of extensive research, the most appropriate methods to measure and localise these signalling radicals are debated and still needs investigation. Many confounding factors such as presence of other reactive intermediates, scavenging enzymes and compartmentation, influence how accurately each can be measured. Further, these signalling radicals have short half-lives ranging from seconds to minutes based on the cellular redox condition. Hence, it is necessary to use the sensitive and specific methods in order to understand the contribution of each signalling molecule to various biological processes. In this review, we will provide current state knowledge on NO measurement in plant samples, via various methods. We will also discuss advantages, limitations and wider applications of each method.
Detection of nitric oxide from chickpea using DAF fluorescence and chemiluminescence methods
(John Wiley & Sons, 2022) Kumari, Aprajita; Bhatoee, Manbir; Singh, Pooja; Kaladhar, Vemula Chandra; Yadav, Nidhi; Paul, Debarati; Loake, Gary J.; Gupta, Kapuganti Jagadis
The free radical nitric oxide (NO) has emerged as an important signal molecule in plants, due to its involvement in various plant growth, development, and stress responses. For elucidating the role of NO, it is very important to precisely determine, localize, and quantify NO levels. Due to a relatively short half-life and its rapid, complex reactivity with other radicals, together with its capacity to diffuse from the source of production, the quantification of NO in whole plants, tissues, organelles, and extracts is notoriously difficult. Hence, it is essential to employ sensitive procedures for precise detection of NO. Currently available methods can fulfill many requirements to precisely determine NO, but each method has several advantages and pitfalls. In this article, we describe a detailed procedure for the measurement of NO by diaminofluorescein (DAF) in cell-permeable forms (DAF-FM-DA). In this method, the tissues are immersed in DAF-FM DA, leading to their diffusion from the plasma membrane to the inside of the cell, where intracellular esterases cleave the ester bonds, leading to DAF-FM release. The resulting DAF-FM reacts with intracellularly generated NO and forms highly fluorescent triazolofluorescein (DAF-FMT), which can be localized and monitored by fluorescence or confocal microscopy, and can also be detected via fluorimetry and flow cytometry. DAF dyes are very popular as they are non-invasive, relatively easy to handle, and commercially available. Another precise and very sensitive method is chemiluminescence detection of NO, where NO reacts with ozone (O3), leading to emission of a quantum of light from which NO can be calculated. Using chickpea seedlings, we describe in detail the measurement of NO using DAF-FM-DA and chemiluminescence methods. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Measurement of nitric oxide from chickpea seedlings using DAF-FM DA fluorescence with fluorescence and confocal microscopy Basic Protocol 2: Chemiluminescence detection of nitric oxide from chickpea seedlings
Differential regulation of nitric oxide mediated by phytoglobin1 plays a role in resistance during Botrytis cinerea infection in Arabidopsis thaliana
(Springer Nature Publishing AG, 2026) Jaiswal, Rekha; Saini, Deepak; Swain, Jagannath; Gupta, Kapuganti Jagadis
Botrytis cinerea is a prominent necrotrophic pathogen responsible for gray mold disease, affecting a wide range of plant species, including economically vital crops such as tomatoes, grapes, strawberries, etc. Nitric oxide (NO) is considered as a crucial player in plant responses to biotic stress. NO homeostasis is regulated by phytoglobin (Pgb1), a potential scavenger of NO. However, the role of the Pgb1-NO cycle in regulating defense response against B. cinerea remains largely unknown. In the current study, we investigated the defense response of Arabidopsis thaliana against B. cinerea infection using antisense (Pgb1 AS) and overexpression (Pgb1 OE) lines, which produce differential levels of NO. The Pgb1 AS line accumulated higher NO levels and conferred resistance against B. cinerea infection, with reduced ROS levels, reduced cell death, and increased stomatal closure. Conversely, Pgb1 OE showed reduced NO levels accompanied by increased susceptibility. The elevated NO level in Pgb1 AS was associated with increased nitrate reductase (NR) activity and upregulation of NIA1 and NIA2 gene expression. Interestingly, ethylene-mediated defense pathway genes such as ERF1, ACS2, and ACS6 were upregulated while pathogen-related genes such as PR1, PR2, PR5, and NPR1 were downregulated in Pgb1 AS line. The elevated expression of ethylene genes corresponded with the higher ethylene levels in Pgb1 AS. Overall, our results confirmed the crucial role of phytoglobin-modulated NO in defense against B. cinerea infection by activating the ethylene-mediated defense pathway.
A discrete role for alternative oxidase under hypoxia to increase nitric oxide and drive energy production
(Elsevier B.V., 2018) Vishwakarma, Abhaypratap; Kumari, Aprajita; Mur, Luis A.J.; Gupta, Kapuganti Jagadis
Alternative oxidase (AOX) is an integral part of the mitochondrial electron transport and can prevent reactive oxygen species (ROS) and nitric oxide (NO) production under non-stressed, normoxic conditions. Here we assessed the roles of AOX by imposing stress under normoxia in comparison to hypoxic conditions using AOX over expressing (AOX OE) and anti-sense (AOX AS) transgenic Arabidopsis seedlings and roots. Under normoxic conditions stress was induced with the defence elicitor flagellin (flg22). AOX OE reduced NO production whilst this was increased in AOX AS. Moreover AOX AS also exhibited an increase in superoxide and therefore peroxynitrite, tyrosine nitration suggesting that scavenging of NO by AOX can prevent toxic peroxynitrite formation under normoxia. In contrast, during hypoxia interestingly we found that AOX is a generator of NO. Thus, the NO produced during hypoxia, was enhanced in AOX OE and suppressed in AOX AS. Additionally, treatment of WT or AOX OE with the AOX inhibitor SHAM inhibited hypoxic NO production. The enhanced levels of NO correlated with expression of non-symbiotic haemoglobin, increased NR activity and ATP production. The ATP generation was suppressed in nia1,2 mutant and non symbiotic haemoglobin antisense line treated with SHAM. Taken together these results suggest that hypoxic NO generation mediated by AOX has a discrete role by feeding into the haemoglobin-NO cycle to drive energy efficiency under conditions of low oxygen tension.
Does the alternative respiratory pathway offer protection against the adverse effects resulting from climate change?
(Oxford University Press, 2020) Florez-Sarasa, Igor; Fernie, Alisdair R; Gupta, Kapuganti Jagadis
Elevated greenhouse gases (GHG) induce adverse conditions directly and indirectly causing decreases in plant productivity. To deal with climate change effects, plants have developed various mechanisms including the fine-tuning of metabolism. Plant respiratory metabolism is highly flexible due to presence of various alternative pathways. The mitochondrial alternative oxidase (AOX) respiratory pathway is responsive to these changes and several lines of evidence suggest it plays a role in reducing excesses of ROS and RNS while providing metabolic flexibility under stress. Here we discuss the importance of the AOX pathway in dealing with elevated carbon dioxide (CO2), nitrogen oxides (NOx), ozone (O3) and the main abiotic stresses induced by climate change. Recent advances in our understanding concerning the in vivo regulation of AOX and its structural properties suggest that novel AOXs with altered regulatory properties could be used in future gene editing strategies. We suggest that fine-tune modulation of the regulatory properties of AOX and targeting its expression in different plant tissues could improve plant growth and productivity under climate change conditions promoted by elevated GHG. Moreover, we also emphasise the need of extensive study on the interactive effects of major global change factors on AOX respiration and the importance of studies differentiating between the roles of AOX in sink versus source tissues under field conditions in order to improve plant productivity in response to elevated GHG.
An efficient method of mitochondrial DNA isolation from Vigna radiata for genomic studies
(Springer Nature Publishing AG, 2020) Singh, Pooja; Sahoo, Ranjan Kumar; Bulle, Mallesham; Gupta, Kapuganti Jagadis
Isolation of mitochondrial DNA from root tissues of mung bean (Vigna radiata) is quite tedious, complex, and often results in low yield. Hence here we show a simple, rapid, and improved protocol for isolation of mitochondrial DNA from root tissues of hydroponically grown mung bean plants. This method involves purification of mitochondria and subsequent isolation of DNA from obtained purified mitochondria. For this purpose, mitochondria were isolated using a discontinuous Percoll gradient centrifugation followed by RNase I treatment to the isolated DNA to remove any traces of RNA contamination. The mitochondrial DNA was isolated from mitochondrial samples by commonly used CTAB method. The specificity of isolated mitochondrial DNA was confirmed using mtDNA-specific genes (NAD1 and COX3). β-Actin primer was used to check the nuclear DNA contamination. PCR amplification was observed in mtDNA specific genes NAD1 and COX3 except nuclear encoded β-actin gene suggesting that mitochondrial DNA was not contaminated by nuclear DNA.
The emerging roles of nitric oxide and its associated scavengers-phytoglobins-in plant symbiotic interactions
(Oxford University Press, 2024) Pathak, Pradeep Kumar; Yadav, Nidhi; Kaladhar, Vemula Chandra; Jaiswal, Rekha; Kumari, Aprajita; Igamberdiev, Abir U.; Loake, Gary J.; Gupta, Kapuganti Jagadis
A key feature in the establishment of symbiosis between plants and microbes is the maintenance of the balance between the production of the small redox-related molecule, nitric oxide (NO), and its cognate scavenging pathways. During the establishment of symbiosis, a transition from a normoxic to a microoxic environment often takes place, triggering the production of NO from nitrite via a reductive production pathway. Plant hemoglobins [phytoglobins (Phytogbs)] are a central tenant of NO scavenging, with NO homeostasis maintained via the Phytogb–NO cycle. While the first plant hemoglobin (leghemoglobin), associated with the symbiotic relationship between leguminous plants and bacterial Rhizobium species, was discovered in 1939, most other plant hemoglobins, identified only in the 1990s, were considered as non-symbiotic. From recent studies, it is becoming evident that the role of Phytogbs1 in the establishment and maintenance of plant–bacterial and plant–fungal symbiosis is also essential in roots. Consequently, the division of plant hemoglobins into symbiotic and non-symbiotic groups becomes less justified. While the main function of Phytogbs1 is related to the regulation of NO levels, participation of these proteins in the establishment of symbiotic relationships between plants and microorganisms represents another important dimension among the other processes in which these key redox-regulatory proteins play a central role.
Expanding roles for S-nitrosylation in the regulation of plant immunity
(Elsevier B.V., 2023) Borrowman, Sam; Gupta, Kapuganti Jagadis; Loake, Gary J.
Following pathogen recognition, plant cells produce a nitrosative burst resulting in a striking increase in nitric oxide (NO), altering the redox state of the cell, which subsequently helps orchestrate a plethora of immune responses. NO is a potent redox cue, efficiently relayed between proteins through its co-valent attachment to highly specific, powerfully reactive protein cysteine (Cys) thiols, resulting in formation of protein S-nitrosothiols (SNOs). This process, known as S-nitrosylation, can modulate the function of target proteins, enabling responsiveness to cellular redox changes. Key targets of S-nitrosylation control the production of reactive oxygen species (ROS), the transcription of immune-response genes, the triggering of the hypersensitive response (HR) and the establishment of systemic acquired resistance (SAR). Here, we bring together recent advances in the control of plant immunity by S-nitrosylation, furthering our appreciation of how changes in cellular redox status reprogramme plant immune function.
Expression analysis of important genes involved in nitrogen metabolism under hypoxia
(Springer Nature Publishing AG, 2020) Bulle, Mallesham; Kishorekumar, Reddy; Wany, Aakanksha; Gupta, Kapuganti Jagadis
Hypoxia or anoxia condition can occurs during flooding or waterlogging and can cause intense damage to the plants. Since oxygen is important for active operation of electron transport chain in mitochondria to generate energy production (ATP) any drop in oxygen can cause an energy crisis during flooding/waterlogging. To cope with this energy crisis plants have developed various anatomical, physiological, and biochemical adaptations. Perception of signals and induction of genes are required for initiation of these adaptive responses. Various genes involved in nitrogen, carbon, and fermentative metabolism play a role in hypoxic tolerance. Regulation of genes involved in nitrogen metabolism also plays a role under hypoxia. Hence in this present chapter we describe the expression of nitrate reductase-1 (NIA1), nitrate reductase-2 (NIA2), and glutamine synthetase-1 (GLN-1) during hypoxia in Arabidopsis.
Expression analysis of ROS-related genes during the germination of chickpea (Cicer Arietinum L.) seeds
(Springer Nature Publishing AG, 2026) Babuta, Priyanka; Samant, Sanjib Bal; Saini, Deepak; Gupta, Kapuganti Jagadis
Seed germination is a critical physiological process that transforms a quiescent seed into a metabolically active seedling and is also a crucial factor in determining maximum crop production. This transition is influenced by various intrinsic and extrinsic factors. Interestingly, reactive oxygen species (ROS) plays an important role in breaking seed dormancy by oxidation of biomolecules, weakening of the testa and degradation of endosperm. Similarly, molecular internal oxygen is also considered vital for the transition of dormancy to seed germination. However, it is essential to establish a correlation between the internal oxygen and the generation of ROS during seed germination. This chapter details protocols for imaging internal oxygen concentrations using VisiSens and fluorescent detection of ROS using H2DCFDA in chickpea seeds, complemented by qPCR analysis of key ROS-related genes (RBOH, AOX 1, UCP 1, and NADH dehydrogenase). These findings from these methods help advance our understanding of the inverse relationship between molecular oxygen and ROS dynamics during seed germination.