Browsing by Author "DEEPIKA KUMAR UMESH"
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ThesisItem Open Access MOLECULAR ANALYSIS OF JASMONIC ACID & POLYAMINES SYNTHESIS & SIGNALING UNDER HEAT & DROUGHT STRESS IN RICE(DIVISION OF PLANT PHYSIOLOGY ICAR-INDIAN AGRICULTURAL RESEARCH INSTITUTE NEW DELHI, 2017) DEEPIKA KUMAR UMESH; Madan Pal SinghRice (OryzasativaL.) is the most important food crop and grown mainly in tropical and subtropical zones.Under current climate change scenarioheat and drought stress have been reported to affect rice productivity. To address the challenges posed by climate change, identifying true sources of heat and drought tolerance and finding reliable ways to ameliorate the effect of these stresses is the need of hour. The present investigation was conducted to test the potential role of polyamines (PAs) and jasmonic acid (JA) in ameliorating the impact of drought and heat stress at anthesis stage in rice. The study was planned with ten genotypes namely N-22, N-L-44, SahbhagiDhan, CR-262-4, Anjali, Vandana, IR-64, PB-1, Swarna and PS-5, raised under normal environment and exposed to drought and heat stresses at anthesisduring 2013-14. Before exposure to stress (at booting stage), the plants were pre-treated with phytohormones JA (1mM) and PAs (a mixture of Put, Spd and Spm with 0.5 mM concentration of each). All the ten genotypes showed reduction in pollen viability (PV), spikelet fertility (SF) and panicle grain weight (PGW) under both the stresses. Four genotypes Swarna, IR-64, PB-1 and PS-5 showed more reductions in above traits and were considered most sensitive to both the stresses. SahbhagiDhan, Vandana, Anjali and CR-262-4 were found moderately tolerant to stress compared to others. N-22 and NL-44 showed least reductions under stress in panicle grain weight and considered as tolerant types. Exogenous application of the PAs and JA showed marginal improvement in PV, SF and PGW.During 2014-2015, four genotypes from above viz. N-22, N-L-44, Swarna and PS-5 were selected based on their performance under above stresses and used for physiological, biochemical and molecular traits like RWC, MSI, H2O2, TBARS, SOD, endogenous level of hormones (PAs and JA) and relative expression of genes involved in synthesis and signalling of these hormones. In general, RWC and MSI decreased and H2O2, TBARS and SOD increased under stress in all genotypes. Reduction in RWC and MSI and increase in H2O2, TBARS were more in sensitive genotypes (Swarna and PS-5) compared to tolerant types (N22 and N-L-44). SOD activity increased more in tolerant genotypes. PV, SF and panicle grain weight gave almost similar results like previous year i.e. all the three traits decreased significantly under both the stresses. Exogenous application of JA and PAs did not show any ameliorative effect on any of the parameters under both the stresses. Under drought condition Put contentof tolerant genotypes N-22 and N-L-44 increased significantly (179.3% and 193.5% respectively), whereas, in Swarna and PS-5 increased by 171.9% and 94.1% respectively. Under heat stress condition increase in Put level was not significant. Under drought condition JA contentof tolerant genotypes N-22 and N-L-44 increased by 57.3% and 85.7%, respectively, whereas, the sensitive genotypes Swarna and PS-5 shown an increase of 42% and 32.7%, respectively. Under heat stress conditionsthere was no significant change in JA levels. Similar trend was observed for relative expression of genes involved in synthesis and signalling of PAs (samdc, adc-1, adc-2 and myb-8) and JA (aos, aoc, opr, coi-1, rsOspr-10 and jaz). This indicates that both PAs and JA may have important role in drought stress alleviation. However, exogenous application of JA and PAs did not show any appreciable changes. One of the reason for insignificant effect of exogenous application of phytohormones could be the lower concentrations applied. Functional validation of Osjazgene was done in Arabidopsis plant. OE-JAZ transgenic plant showed more expression of Osjazgene compared to WT (Wild Type) plants under stress. Osjazis a repressor of JA signalling and we found that OE-JAZ transgenic linesshowed less increase in endogenous level of JA compared to WT under drought conditions. Findings of this study conclude that pre-treatment with 0.5 mM concentration of PAs and 1.0 mM concentration of JA was not effective to ameliorate the effects of drought and heat stress effect on PV, SF and PGW. But both the growth regulators lowered the level of oxidative stress induced by drought and heat stress, but the magnitude of impact was not effective to show changes in PV, SF and PGW. It indicates that the changes in the endogenous level of PAs and JA due to application of both the growth regulators were less than the threshold levels required to ameliorate the effect of heat and drought stress. The study may be further undertaken in future with different and higher concentrations of phyto-hormones which could enhance the endogenous levels up to reach the threshold levels for their amelioration effects under heat and drought stress in riceThesisItem Open Access PROTEIN OXIDATION IN RESPONSE TO NITROGEN STRESS IN WHEAT SEEDLINGS(IARI, DIVISION OF PLANT PHYSIOLOGY, 2011) DEEPIKA KUMAR UMESH; Vanita JainNitrogen is one of the most important nutrients required by plants and its deficiency is also widespread. Sixty percent of the soils in India are deficient in nitrogen (N). This deficiency of N seriously affects plant growth, yield and quality. Lot of work in diverse areas has been done but many questions regarding N nutrition remain unanswered. The nitrogen in wheat is taken up mainly in the form of nitrate, and occasionally in the form of ammonium. But, wheat plants although capable of taking up ammonium are sensitive to ammonium nutrition. In terms of quality the protein content goes down. When plant faces nitrogen stress, it is possible that due to stress in spite of low protein content of tissues, they might be oxidised and carbonylated leading to increased degradation in such plants facing nutrient stress. In the present study the, the effect of different levels and forms of N has been investigated in wheat seedlings grown under controlled conditions. The interaction between cytokinins gene expression and N was also investigated. The extent of protein carbonylation was analyzed and an attempt was made to elucidate the pathway of protein degradation. Wheat seedlings (var. PBW 343) were grown under four different N treatments and various parameters of growth, N-assimilation and stress was studied. Result indicated that growth and nitrogen metabolism was enhanced in seedlings under high NO3- -N when compared to those grown with low and without N. The growth of the wheat seedlings was severely inhibited in terms of biomass accumulation, leaf area and root: shoot ratio was enhanced in zero N and ammonium grown plants as compared to the nitrate N fed wheat seedlings. There was almost 5 fold increase in shoot dry weight of NO3- -N fed seedlings when compared to those grown without N. In seedling grown with 5mM NH4+-N shoot dry weight declined by 63% as compared to the wheat seedlings grown in 5 mM NO3- -N. There was a relative increase in shoot length by 20% to 40% in the seedlings grown in solution having N salts as compared to those grown without N. In zero-N and ammonium fed plants the nitrate levels, total reduced N and total soluble protein content was significantly lower as compared to both low and high NO3- -N grown wheat seedlings. These parameters directly correlated with low NR activity in zero-N and NH4+-N treatments. There was increased production of ROS in terms of increase in superoxide radicals and H2O2 in NH4+-N fed wheat seedlings followed by those grown without N. Increase in protease activity, total free amino acids and TBARS was also observed in these treatments. Under normal conditions, plants contain numerous carbonylated proteins, which are thought to be indicative of oxidative stress damage. Conditions that promote formation of reactive oxygen species (ROS) enhance protein carbonylation, and protein degradation is required to reverse the damage. However, it is not clear how the degradation of carbonylated proteins is controlled in planta. In this report, we show that detached wheat leaves rapidly and selectively degrade carbonylated proteins when kept in the dark as compared to the detached leaves incubated in light or in the intact seedlings. The loss of carbonylated proteins corresponded to a loss of soluble protein and accumulation of free amino acids. Degradation of carbonylated proteins was blocked by carbobenzoxyl-leucinyl-leucinyl-leucinal (MG132) in dark detached leaves, but not by 3-methyladenine, suggesting that the 26S proteasome pathway rather than the autophagic pathway was involved in degrading protein carbonyls in dark detached leaves. In light incubation pathway the use of inhibitors revealed that both autophagic and proteasomal pathways were involved in degrading protein carbonyls. The study also indicated the role of cytokinins in preventing the the protein carbonyl degradation. The expression of IPT3 gene of cytokinin biosynthetic pathway was almost absent in the seedlings grown at zero N and NH4+-N, and in these treatments the degradation of protein carbonyls was high as supported by low level of total soluble proteins and high levels of free amino acids. Exogenous