| Nitrate (NO3-) and ammonium (NH4+) are the two major nitrogen (N) sources taken up from the soil by higher plants. Previous results showed that, appropriate ratio of NO3- to NH4+ could promote the growth of tomato. With the increase of NO3-proportion to NH4+ in the solution, it was found that large amounts of citric acid and malic acid were accumulated in the leaves, indicating that the metabolism of NO3- to NH4+ in the plant was closely related to the metabolism of organic acid. Although there have been some reports on the dynamic changes in the activity of related enzyme involved in the metabolism of organci acid during the development of fruit, it was still unclear that how the different N forms affected the metabolism of organic acid in tomato fruit in physiological and molecular level. Thus, in this experiment, cherry tomato "Red Virgin" was used to study the pathway of organic acid produced in the leaves, and then dynamic changes of the activity of enzymes involved in nitrogen assimilation, organic acid metabolism and the expression of related genes during the development of fruit. The results are as follows:1. The 14C-malic acid and 14C- citric acid synthesized in leaves was only transported to the petiole within 1h.6h and 17h after 14CO2 tags, only the radioactivity of 14C-malic acid was detected in tomato roots, while no detection of 14C-citric acid.14C-malic acid detected in the fruit placenta was not transported from the leaves, however, it was transformed by the carbohydrate which was previously transported to the fruits.2. The single fruit weight was slightly higher under 75% NO3-:25% NH4+ treatment than that under NO3- treatment, also the levels of NH4+, total amino acid, total nitrogen content and nitrogen accumulation in fruit. The results here suggested that combination of NO3- with NH4+ could promote the nitrogen metabolism in fruit. In comparison with 100% NH4+ treatment, both treatments of 75% NO3-:25% NH4+ and 100% NO3 led to higher NR activity and gene expression, but there is no significant difference between treatments of 75% NO3-:25% NH4+ and 100% NO3-. The treatment of 75% NO3-:25% NH4+ showed significant higher GS activity than that under treatment of 100%NO3-, which was also correlated to the NH4+content. It seemed that the GS activity was related to the NH4+supply.The expression of isozyme GS1 (Cytosolic type GS) and GS2 (Chloroplast type GS) was inconsistent with GS activity under different nitrogen forms. It was indicated that GS activity might be also regulated at posttranscriptional level. These results suggest that the impact of different N forms on NR and GS activity and expression might result great differently in N metabolism in tomato fruit.3. The citrate content was significantly higher than that of malate in the latter developmental stage. compared with 100% NO3-,25% NH4+ replacement of NO3- led to a significant lower contents in both citrate and malate at mature stage. The citrate content and the activity of CS showed a similar change tendency as a single peak during the development of fruit, however, they peaked at different stage. The content of malate supplied by 100% NH4+ displayed a significant positive correlationship with PEPC activity, and the content of malate and citrate supplied by 100% NO3- and combined 75%NO3- with 25% NH4+ showed a significant positive correlationship with MDH activiy. At the enlargement stage and mature stage, the expression of PEPC gene (PEPC1 and PEPC2) supplied by 100% NO3- and combined 75% NO3-with 25% NH4+ was both significantly higher than that of 100% NH4+, which showed a consistent change tendency of PEPC activiy and the content of malate and citrate. The expression of mitochondrial MDH (mMDH) was corresponding to the activity of MDH and the content of malate, especially in the supply of 100% NO3-, which indicating that mMDH might play a major role in the activiy of MDH and malate accumulation in fruit. The expression of CS gene also showed a consistent tendency with the activity of CS. |
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