Nickel Stress Response And Nitric Oxide/Hydrogen Sulfide-induced Nickel Tolerant Mechanisms In Rice (oryza Sativa L.) Seedlings

Nickel?Ni?pollution has been reported across the world,including Asia,Europe and North America and threatens the health of vegetation,wildlife,and human beings.China is one of the leading extractors and manufacturers of Ni,and the consumers of Ni products.Recently,a joint report on the status of soil contamination in China issued by Chinese Ministry of Environmental Protection?MEP?and Ministry of Land and Resources?MLR?mentioned that Ni contamination comprised more than 4.8% out of 19.4% total contamination in Chinese arable lands.Thus,Ni contamination of arable soil in China is one of the utmost environmental problem and understanding of Ni toxicity in rice is of great significance in safety production of rice and remediation of Ni polluted paddy soils.However,there are few researches on the effects of Ni toxicity and detoxicity as well as the deeper mechanisms in rice.Here,a series of hydroponic experiments were carried out in controlled conditions to unravel the physiological,biochemical,metabolic and molecular responses in Ni-stressed rice seedlings in the presence and absence of NO and H₂ S,the notable biological messengers in plant tissues to regulate numerous growth and development processes.It was hypothesized that exogenous application of NO and H₂ S donors might have a potential to confer tolerance to Ni stress by regulating various metabolic processes,modulating the production of osmolytes,enhanced antioxidative defense system at activity as well as genetic level.The main findings of these studies are summarized below.1.The effects of Ni exposure on the rice seedlings were investigated.The results showed that high Ni concentrations in the growth medium resulted a decline in root and shoot lengths,fresh weight?FW?and dry weight?DW?of rice plants,which relating to the depletion of the contents of photosynthetic pigments.Measurement of Ni concentrations in the roots and shoots showed that Ni was mainly accumulated in roots followed by shoots.Moreover,Ni was mainly deposited in soluble fraction and cell wall,then cell organelle,which suggests that both compartments acts as crucial defensive barriers against Ni toxicity in rice plants.Ni also induced its toxicity by damaging oxidative metabolism,as indicated by increased level of hydrogen peroxide?H₂O2?and malondialdehyde?MDA?content.Furthermore,Ni stress also showed a desynchronized antioxidant system by increasing the activities of catalase?CAT?,peroxidase?POD?and the contents of ascorbic acid?As A?and glutathione?GSH?,whereas decreasing the activity of superoxide dismutase?SOD?in the roots and shoots of rice plants.Ni stress also triggered the rate of proline accumulation and decreasing the contents of soluble protein and soluble sugar.2.Studies on the effect of Ni stress on nitrogen metabolism in rice showed that Ni exposure of rice plants resulted decline in nitrate?NO3-?concentration in leaves,with the increase of Ni application in growth medium.At highest Ni concentration?200 ?M?,nitrate reductase?NR?,nitrite reductase?Ni R?,glutamine synthetase?GS?and glutamate synthase?GOGAT?activities were significantly declined by 56%,39%,44% and 11%,respectively,over control.In contrast,ammonium?NH₄⁺?concentration and the activity of glutamate dehydrogenase?GDH?increased sharply by 48% and 46%,respectively,as compared with control.Furthermore,most active aminotransferases,such as glutamic-oxaloacetic transaminase?GOT?and glutamic-pyruvic transaminase?GPT?activity were also inhibited by 48% and 36%,respectively,in comparison with control.3.Application of exogenous sodium nitroprusside?SNP?,a NO donor,in rice grown under excessive Ni significantly improved the growth of rice seedlings.The enhanced Ni-tolerance of rice prompted by SNP could be ascribed to its ability to regulate Ni uptake,decrease Ni-induced oxidative stress as evidenced by reduced levels of H₂O2,MDA,and electrolyte leakage in Ni-stressed plants.The positive roles of NO against Ni-toxicity also reflected through its protective effects on photosynthetic pigments,soluble proteins and proline.SNP also boosted antioxidant capacity in Ni-stressed plants by maintaining increased levels of As A,enhanced activities of ROS-detoxifying enzymes,particularly p POD)and CAT in both roots and shoots compared with Ni-stressed alone plants.Moreover,SNP treatment also upregulated the transcript levels of genes of CAT,POD,ascorbate peroxidase,glutathione reductase and superoxide dismutase in shoots under Ni-stress.Using different sulfide compounds and NO scavenger c PTIO,we also provided evidence that NO,rather than other byproducts of SNP,contributed to the improved performance of rice seedlings under Ni-stress.4.Exogenous application of sodium hydrosulfide?Na HS?,H₂ S donor successfully reversed the adverse effects of Ni,as reflected in plant growth,photosynthesis,stomatal conductance,transpiration and internal CO2 concentration.Transmission electron microscopy of chloroplast ultrastructure clearly revealed a protective role of H₂ S on photosynthetic machinery of rice,thus contributing efficient photosynthesis upon Ni stress.Na HS also restricted the uptake and accumulation of Ni,as evidenced by reduced content of Ni in roots and leaves of Na HS-supplemented Ni-stressed plants.Na HS reduced Ni-induced oxidative stress,particularly by enhancing As A,GSH content,and the activities of reactive oxygen species detoxifying enzymes.A rescue of Ni-caused decrease in NO₃^- content and the activities NO₃^- biosynthesizing enzymes NR,Ni R,GS,GOGAT,GOT and GPT in leaves indicated a positive role of H₂ S on NO₃^- metabolism in rice under Ni stress.Na HS application also revered Ni-mediated increases in NH₄⁺ content and GDH activity,implying H₂S-induced alleviation of NH₄⁺ toxicity.The regulatory response of H₂ S on nitrogen metabolism was further confirmed by increased and decreased transcript abundance of NO₃^- and NH₄⁺ metabolism associated genes,respectively.