The role of nitrate reductase and asparagine synthetase in soybean (Glycine max L.) under nitrogen stress / Papel da redutase do nitrato e da asparagina sintetase em plantas de soja (Glycine max L.) sob condições de estresse de nitrogenio

AUTOR(ES)
DATA DE PUBLICAÇÃO

2007

RESUMO

The root system of soybean (Glycine max L.) is an important site for the assimilation of nitrogen, whether by nitrate assimilation in the roots, or by symbiotic nitrogen fixation in the nodules. The main products of inorganic nitrogen assimilation, the amino acids asparagine and glutamine and the ureides allantoin and allantoic acid, are used in the transport of assimilated nitrogen to the shoot. Thereby, these products represent a source of reduced nitrogen for the sink tissues, for the formation of other amino acids, proteins, nucleic acids and all the other nitrogenous compounds synthesized in the cell. Therefore, the objective of this study was to relate changes in the transport of nitrogen in the xylem of soybean caused by nitrogen deficiency with the behaviour of certain enzymes of nitrogen assimilation in the root system. Nitrogen deficiency was induced by the transfer of soybean plants to a hydroponic system without nitrate or any other source of inorganic nitrogen. In the case of non-nodulated plants deficiency was imposed by the interruption of nitrate assimilation by the roots, and for the nodulated plants nitrogen fixation was inhibited by immersion of the nodules in the hydroponic system. Under nitrogen deficiency, an increase in the aspartate/asparagine (ASP/ASN) ratio of the xylem sap was observed in both nodulated and non-nodulated plants. Nevertheless, this increase was substantial only on the first day of treatment. The recovery from the stress was only observed for the non-nodulated plants, where the levels of ASP and ASN returned to their initial values. Analyses of asparagine synthetase expression in the root system of soybean during treatment and recovery from nitrogen stress indicates that this enzyme can underlay the changes in ASP/ASN ratios. AS is an enzyme that catalyses the biosynthesis of Asn, by transferring the amide nitrogen from glutamine to aspartate, producing asparagine and glutamate. The fall in N supply, whether by interruption of nitrate assimilation or nitrogen fixation, leads to a decline in glutamine, the immediate product of ammonium ion assimilation via glutamine synthetase. With the reduction in glutamine levels the activity of AS is also reduced resulting in diminished utilization of ASP. In non-nodulated plants, after the transfer to the hydroponic system without nitrate, the expression of the three genes declines sharply, and after the return of the plants to a supply of nitrate the AS genes are expression again. In nodulated plants the expression of AS was also reduced during treatment, however, in this case only the gene SAS1 was affected. The SAS1 gene did not recover its initial levels of expression after removing the stress which is further evidence for the correlation between AS activity and the changes in ASP/ASN ratios in the xylem sap, since these ratios were also not recovered. In view of the importance of nitrate reductase (NR) in the process of nitrate assimilation by the root system, its behaviour was evaluated during nitrate deficiency and recovery. The activity of NR increased considerably when plants were supplied with nitrate, in the case of roots of non-nodulated as well as the root system (roots and nodules) of nodulated plants. When non-nodulated plants were transferred to a nutrient solution free of nitrate, NR activity fell sharply, almost disappearing within 24 hours. RN activities in roots and nodules of soybean grown in the absence of nitrate were very low, with activity in the nodules being somewhat higher than in the roots. The low activity found in the roots and nodules could be due to a constitutive enzyme since plants were grown throughout with nitrate-free medium. However, the possible presence of trace amounts of nitrate in the tap water used to irrigate the plants cannot be discarded and may have been sufficient to induce the low levels of enzyme found. If on the one hand there are doubts as to the presence of a constitutive enzyme, the presence of the induced form is very clear. In any case, the strong dependence of NR on the presence of nitrate is an unknown fact for soybean roots. Nitrate also affected the expression of AS in roots of nodulated plants grown without any mineral source of nitrogen, since its expression increased tremendously when such plants were supplied with nitrate. The data concerning the behaviour of the enzymes studied here indicate that alterations in xylem amino acids of soybean plants subjected to nitrogen deficiency are related to the assimilatory processes of the root system, and that AS appears to be mainly responsible for the changes in ASP/ASN ratios

ASSUNTO(S)

xylem xilema nitrato redutases nitrogenio - metabolismo asparaginase nitrogen soybean asparaginase soja nitrate reductases

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