Publications of NIPGR Scientists
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Item 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 JagadisA 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.Item Nitric oxide regulates mitochondrial biogenesis in plants(John Wiley & Sons, 2023) Kumari, Aprajita; Kaladhar, Vemula Chandra; Yadav, Nidhi; Singh, Pooja; Reddy, Kishorekumar; Gupta, Kapuganti JagadisThe site of nitric oxide (NO) production in mitochondrial cytochrome c oxidase and the role of NO in mitochondrial biogenesis are not known in plants. By imposing osmotic stress and recovery on Arabidopsis seedlings we investigated the site of NO production and its role in mitochondrial biogenesis. Osmotic stress reduced growth and mitochondrial number while increasing NO production. During the recovery phase the mitochondrial number increased and this increase was higher in wild type and the high NO-producing Pgb1 silencing line in comparison to the NO-deficient nitrate reductase double mutant (nia1/nia2). Application of nitrite stimulated NO production and mitochondrial number in the nia1/nia2 mutant. Osmotic stress induced COX6b−3 and COA6-L genes encoding subunits of COX. The mutants cox6b-3 and coa6-l were impaired both in NO production and mitochondrial number during stress to recovery suggesting the involvement of these subunits in nitrite-dependent NO production. Transcripts encoding the mitochondrial protein import machinery showed reduced expression in cox6b-3 and coa6-l mutants. Finally, COX6b-3 and COA6-L interacted with the VQ27 motif-containing protein in the presence of NO. The vq27 mutant was impaired in mitochondrial biogenesis. Our results suggest the involvement of COX derived NO in mitochondrial biogenesis.Item Phytoglobin-NO cycle and AOX pathway play a role in anaerobic germination and growth of deepwater rice(John Wiley & Sons, 2021) Kumari, Aprajita; Singh, Pooja; Kaladhar, Vemula Chandra; Manbir; Paul, Debarati; Pathak, Pradeep Kumar; Gupta, Kapuganti JagadisAn important and interesting feature of rice is that it can germinate under anoxic conditions. Though several biochemical adaptive mechanisms play an important role in the anaerobic germination of rice but the role of phytoglobin-nitric oxide cycle and alternative oxidase pathway is not known, therefore in this study we investigated the role of these pathways in anaerobic germination. Under anoxic conditions, deepwater rice germinated much higher and rapidly than aerobic condition and the anaerobic germination and growth were much higher in the presence of nitrite. The addition of nitrite stimulated NR activity and NO production. Important components of phytoglobin-NO cycle such as methaemoglobin reductase activity, expression of Phytoglobin1, NIA1 were elevated under anaerobic conditions in the presence of nitrite. The operation of phytoglobin-NO cycle also enhanced anaerobic ATP generation, LDH, ADH activities and in parallel ethylene levels were also enhanced. Interestingly nitrite suppressed the ROS production and lipid peroxidation. The reduction of ROS was accompanied by enhanced expression of mitochondrial alternative oxidase protein and its capacity. Application of AOX inhibitor SHAM inhibited the anoxic growth mediated by nitrite. In addition, nitrite improved the submergence tolerance of seedlings. Our study revealed that nitrite driven phytoglobin-NO cycle and AOX are crucial players in anaerobic germination and growth of deepwater rice.Item The power of the phytoglobin-NO cycle in the regulation of nodulation and symbiotic nitrogen fixation(John Wiley & Sons, Inc., 2020) Singh, Pooja; Kumari, Aprajita; Foyer, Christine H.; Gupta, Kapuganti JagadisRecent years have seen a large increase in our understanding of the multifaceted roles of nitric oxide (NO) in plant biology, particularly in the regulation of symbiotic interactions with soil microorganisms. NO serves key functions in the initiation and maintenance of legume–rhizobium symbiosis. Extensive molecular crosstalk between the host legume and the soil rhizobium results in the initiation of symbiosis and the establishment of a highly specialized organ called the nodule, which provides the conditions required for bacterial nitrogen fixation. Within the nodules, the rhizobium bacteria are housed in bacteroids, in which the oxygen is maintained at very low levels allowing the bacterial nitrogenase to function. This enzyme is oxygen sensitive (half maximal inhibitory concentration (IC50) oxygen for nitrogenase is 2.9 µM; Goldberg et al., 1987). Hence, a key feature of the nodule is the maintenance of a strong oxygen diffusion barrier, in which leghaemoglobin is used to transfer oxygen to the plant mitochondria to sustain respiration.Item Nitrite and nitric oxide are important in the adjustment of primary metabolism during the hypersensitive response in tobacco(Oxford University Press, 2019) Mur, Luis A. J.; Kumari, Aprajita; Brotman, Yariv; Zeier, Jurgen; Mandon, Julien; Cristescu, Simona M.; Harren, Frans; Kaiser, Werner M.; Fernie, Alisdair R.; Gupta, Kapuganti JagadisNitrate and ammonia deferentially modulate primary metabolism during the hypersensitive response in tobacco. In this study, tobacco RNAi lines with low nitrite reductase (NiRr) levels were used to investigate the roles of nitrite and nitric oxide (NO) in this process. The lines accumulate NO2-, with increased NO generation, but allow sufficient reduction to NH4+ to maintain plant viability. For wild-type (WT) and NiRr plants grown with NO3-, inoculation with the non-host biotrophic pathogen Pseudomonas syringae pv. phaseolicola induced an accumulation of nitrite and NO, together with a hypersensitive response (HR) that resulted in decreased bacterial growth, increased electrolyte leakage, and enhanced pathogen resistance gene expression. These responses were greater with increases in NO or NO2- levels in NiRr plants than in the WT under NO3- nutrition. In contrast, WT and NiRr plants grown with NH4+ exhibited compromised resistance. A metabolomic analysis detected 141 metabolites whose abundance was differentially changed as a result of exposure to the pathogen and in response to accumulation of NO or NO2-. Of these, 13 were involved in primary metabolism and most were linked to amino acid and energy metabolism. HR-associated changes in metabolism that are often linked with primary nitrate assimilation may therefore be influenced by nitrite and NO production.Item Nitric oxide accelerates germination via the regulation of respiration in chickpea(Oxford University Press, 2019) Pandey, Sonika; Kumari, Aprajita; Shree, Manu; Kumar, Vinod; Singh, Pooja; Bharadwaj, Chellapilla; Loake, Gary J.; Parida, Swarup K.; Masakapalli, Shyam Kumar; Gupta, Kapuganti JagadisSeed germination is crucial for the plant life cycle. We investigated the role of nitric oxide (NO) in two chickpea varieties that differ in germination capacity: Kabuli, which has a low rate of germination and germinates slowly, and Desi, which shows improved germination properties. Desi produced more NO than Kabuli and had lower respiratory rates. As a result of the high respiration rates, Kabuli had higher levels of reactive oxygen species (ROS). Treatment with the NO donor S-nitroso-N-acetyl-D,L-penicillamine (SNAP) reduced respiration in Kabuli and decreased ROS levels, resulting in accelerated germination rates. These findings suggest that NO plays a key role in the germination of Kabuli. SNAP increased the levels of transcripts encoding enzymes involved in carbohydrate metabolism and the cell cycle. Moreover, the levels of amino acids and organic acids were increased in Kabuli as a result of SNAP treatment. 1 H-nuclear magnetic resonance analysis revealed that Kabuli has a higher capacity for glucose oxidation than Desi. An observed SNAP-induced increase in 13C incorporation into soluble alanine may result from enhanced oxidation of exogenous [13C]glucose via glycolysis and the pentose phosphate pathway. A homozygous hybrid that originated from a recombinant inbred line population of a cross between Desi and Kabuli germinated faster and had increased NO levels and a reduced accumulation of ROS compared with Kabuli. Taken together, these findings demonstrate the importance of NO in chickpea germination via the control of respiration and ROS accumulation.Item The role of nitrite and nitric oxide under low oxygen conditions in plants(Oxford University Press, 2020) Gupta, Kapuganti Jagadis; Mur, Luis A.J.; Wany, Aakanksha; Kumari, Aprajita; Fernie, Alisdair R.; Ratcliffe, R. GeorgePlant tissues, particularly roots, can be subjected to periods of hypoxia due to environmental circumstances. Plants have developed various adaptations in response to hypoxic stress and these have been extensively described. Less well-appreciated is the body of evidence demonstrating that scavenging of nitric oxide (NO) and the reduction of nitrate/nitrite regulate important mechanisms that contribute to tolerance to hypoxia. Whilst ethylene controls hyponasty and aerenchyma formation, NO production apparently regulates hypoxic ethylene biosynthesis. In the hypoxic mitochondrion, cytochrome c oxidase, which is a major source of NO, is also inhibited by NO, thereby reducing the respiratory rate and enhancing local oxygen concentrations. Nitrite can maintain ATP generation under hypoxia by coupling its reduction to the translocation of protons from the inner side of mitochondria and generating an electrochemical gradient. This reaction can be further coupled to a reaction whereby non-symbiotic haemoglobin oxidizes NO to nitrate. In addition to these functions, nitrite has been reported to influence mitochondrial structure and supercomplex formation, as well as playing a role in oxygen sensing via the N-end rule pathway. These studies establish that nitrite and NO perform multiple functions during plant hypoxia and suggest that further research into the underlying mechanisms is warranted.Item 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 JagadisPlant 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.Item Pathways of nitric oxide metabolism and operation of phytoglobins in legume nodules: missing links and future directions(John Wiley & Sons, 2018) Berger, Antoine; Brouquisse, Renaud; Pathak, Pradeep Kumar; Hichri, Imène; Singh, Inderjit; Bhatia, Sabhyata; Boscari, Alexandre; Igamberdiev, Abir U.; Gupta, Kapuganti JagadisThe interaction between legumes and rhizobia leads to the establishment of a beneficial symbiotic relationship. Recent advances in legume - rhizobium symbiosis revealed that various reactive oxygen and nitrogen species including nitric oxide (NO) play important roles during this process. Nodule development occurs with a transition from a normoxic environment during the establishment of symbiosis to a microoxic environment in functional nodules. Such oxygen dynamics are required for activation and repression of various NO production and scavenging pathways. Both the plant and bacterial partners participate in the synthesis and degradation of NO. However, the pathways of NO production and degradation as well as their cross-talk and involvement in the metabolism are still a matter of debate. The plant-originated reductive pathways are known to contribute to the NO production in nodules under hypoxic conditions. Non-symbiotic hemoglobin (phytoglobin) (Pgb) possesses high NO oxygenation capacity, buffers and scavenges NO. Its operation, through a respiratory cycle called Pgb-NO cycle, leads to the maintenance of redox and energy balance in nodules. The role of Pgb/NO cycle under fluctuating NO production from soil needs further investigation for complete understanding of NO regulatory mechanism governing nodule development to attain optimal food security under changing environment.Item Nitric oxide is essential for the development of aerenchyma in wheat roots under hypoxic stress(John Wiley & Sons, 2017) Wany, Aakanksha; Kumari, Aprajita; Gupta, Kapuganti JagadisIn response to flooding/waterlogging, plants develop various anatomical changes including the formation of lysigenous aerenchyma for the delivery of oxygen to roots. Under hypoxia, plants produce high levels of nitric oxide (NO) but the role of this molecule in plant-adaptive response to hypoxia is not known. Here, we investigated whether ethylene-induced aerenchyma requires hypoxia-induced NO. Under hypoxic conditions, wheat roots produced NO apparently via nitrate reductase and scavenging of NO led to a marked reduction in aerenchyma formation. Interestingly, we found that hypoxically induced NO is important for induction of the ethylene biosynthetic genes encoding ACC synthase and ACC oxidase. Hypoxia-induced NO accelerated production of reactive oxygen species, lipid peroxidation, and protein tyrosine nitration. Other events related to cell death such as increased conductivity, increased cellulase activity, DNA fragmentation, and cytoplasmic streaming occurred under hypoxia, and opposing effects were observed by scavenging NO. The NO scavenger cPTIO (2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide potassium salt) and ethylene biosynthetic inhibitor CoCl2 both led to reduced induction of genes involved in signal transduction such as phospholipase C, G protein alpha subunit, calcium-dependent protein kinase family genes CDPK, CDPK2, CDPK 4, Ca-CAMK, inositol 1,4,5-trisphosphate 5-phosphatase 1, and protein kinase suggesting that hypoxically induced NO is essential for the development of aerenchyma.
