Effects Of Nitrogen Fertilizer Form And Dosage On Cadmium Accumulation And Toxicity And Its Regulatory Mechanism In Rice

Cadmium（Cd） is regarded as a toxic heavy metal for plants and animals. Cd has been brought into soil mainly by activities of industrial production, mining, sewage irrigation, inferior phosphate fertilizer application and atmospheric precipitation. Cd may easily enter into plant and cause serious problems such as metabolic disorder, nutrition deficiency, chlorophyll synthesis inhibition, photosynthesis reduction, oxidative stress etc, leading to plant growth suppression and even to death. Even more important, accumulation and enrichment of Cd in food chain potentially cause serious problems to human body health. Rice is a main route of non-occupational exposure especially in countries of China, Japan and South Korea. However, the critical value of Cd content in rice is below 0.1 mg Cd/kg dry weight which was permitted by FAO. However, Cd enrichment exceeded in rice has been worried by far more people and there is an urgent need to lower Cd accumulation in rice. Researchers have proposed several approaches to minimize rice Cd absorption and accumulation. In conclusion, reports suggested that nutrition management was considered to be the most economical, time-save, effective and interfere less with crop production to minimize Cd accumulation in rice. However, nitrogen（N） as one of the most significant macro-nutrient elements, played crucial roles in Cd uptake, translocation, accumulation and toxic alleviation in rice, has not been studied clearly and still need to carry out further researches.These studies were carried out with hydroponic and soil cultivation ways. Rice（O. sativa L., cv. Nipponbare） and Yoshida’s rice nutrient solution were used. Choosed standard N concentration of nitrate（NO₃^-）, ammonium（NH₄⁺） and Urea did the only N resource respectively. Cd concentration was set as 100 μM. Effects of Cd uptake, accumulation and toxic alleviation responding to different N forms and dosages were carried out at seedling stage in rice. Results showed that:1. The rice treated with 3 different N fertilizer forms showed different Cd toxicity and Cd accumulation under Cd stress. Plantlets treated with NH₄⁺ showed higher growth ability with higher tillers, dry weight, photosynthetic rate, transpiration rate and total N content by 49.1%, 9.6%, 106.5%, 9.6% and 48.3% when compared to NO₃^- treated plantlets under Cd stress, respectively. Root and shoot malondialdehyde（MDA） content and hydrogen peroxide（H2O2） content significantly lower 44.2%, 18.9%, 48.5% and 18.9% than NO₃^- treated plantlets under Cd stress. Result suggests that plantlet treated with NH₄⁺ significantly reduced Cd toxicity, and the ability was higher than Urea and NO₃^-. Effects of N forms on rice Cd absorption and accumulation were also differently, NO₃^- treated rice plantlet had strongest Cd absorption, which was 2.79 times than NH₄⁺ treated in rice root, followed by Urea and finally was the NH₄⁺ treated plant.2. Excessive NO₃^- enhances Cd uptake by up-regulating Os IRT1 expression in rice. Using hydroponics and soil cultivation, 3 different concentration of NO₃^- fertilizer were applied in rice. The results showed that excessive NO₃^- enhanced Cd uptake by increasing net Cd2+ influx at elongation area in rice root, it was 53.2% higher than control. Therefore, under excessive NO₃^- condition, the content of Cd in rice root, shoot and milled rice were increased by 8%, and 67% and 93%, respectively. On contrary, Cd2+ influx and Cd content sharply decreased under NO₃^- deficiency. Results also showed that excessive NO₃^- up-regulated the iron（Fe） transporter system of Os IRT1 gene expression. Simultaneously, Fe content significantly increased 13.5% and 59.9% in rice root and shoot regardless of Cd. In conclusion, the results suggested that excessive NO₃^- fertilizer application in Cd polluted paddy areas should be avoided to reduce Cd risk in rice.3. Excessive NH₄⁺ alleviated Cd toxicity by increasing PCs content in root and activating antioxidant enzyme activity in rice. Under hydroponic conditions, different concentrations of NH₄⁺ treatment were carried out on 3 week-old rice plantlet（Nipponbare）. The results showed that excessive NH₄⁺ significantly reduced Cd toxicity in rice. The plantlets treated with excessive NH₄⁺ increased plant height, chlorophyll a content, chlorophyll b content, Photosynthetic rate and shoot N concentration by 4.7%, 17.8%, 27.1%, 28.0% and 27.1% under Cd stress, respectively. And NH₄⁺ deficiency significantly decreased these indicators. Furthermore, studies showed that excessive NH₄⁺ enhanced surphur（S） assimilation, S content and S accumulation was increased by 17.8% and 17.4% in rice root respectively under Cd stress. Simultaneous, phytochelatin（PCs） content also significantly elevated by 6.8% in rice root. Furthermore, excessive NH₄⁺ significantly reduced root MDA content, shoot MDA content and root H2O2 content by 13.4%, 13.2% and 23.9%, respectively. NH₄⁺ deficiency significantly increased these indicators. Excessive NH₄⁺ significantly increased relative expression of root Os MT-1 gene and shoot Os LCD gene in rice. Results also showed that excessive NH₄⁺ increased peroxidase（POD） and ascorbic acid oxidase（APX） activity regardless of Cd presence or not, the root and shoot POD and shoot APX activity was increased by 31.3%, 35.3% and 31.3% respectively under Cd stress. All these results indicate that excessive NH₄⁺ reduced Cd toxicity by increasing POD and APX activity.4. Excessive NH₄⁺ significantly increased the ability of Cd translocation from root to shoot. Results showed that the ratio of shoot/root Cd content under excessive NH₄⁺ condition was higher 14.5% than control, and the ratio significantly decreased under NH₄⁺ deficiency. In addition, excessive NH₄⁺ significantly down-regulated Os HMA3 gene expression in rice roots regardless of Cd presence or not, which reduced PCs-Cd fixation in root cell vacuole. Therefore, result indicates that excessive NH₄⁺ enhanced Cd translocation by inhibiting root Os HMA3 gene expression in rice.5. Excessive Urea up-regulating gene expression to enhance Cd absorption in rice. Treated with different dosages of Urea in rice, results showed that excessive Urea increased NO₃^- and NO₃^-/ NH₄⁺ ratio, up-regulated Os Nramp1, Os Nramp5 and Os HMA2 relative expression, and increased root and shoot Cd content in rice, but Urea deficiency down-regulated these indicators.The researches of Cd absorption, accumulation and toxicity responding to N fertilizer, has important theoretical and practical significance for rice production. In our researches, 3 most commonly used N fertilizers were used to study the effect of N on Cd absorption, accumulation and toxicity in rice. Not only provide a theoretical basis to reduce plant Cd absorption, but also provide references on N fertilizer application in Cd pollution paddy soil.