Regulatory Mechanisms Of Nitrogen Metabolism Under Alkali Stress In Rice (Oryza Sativa L.)

Soil alkalization frequently causes severe problems in some areas. Alkali stress is an important agricultural contaminant and has complex effects on plant metabolism specifically root physiology. However, relatively little attention has been given to this problem, and the insight into mechanisms of alkali tolerance was lacking. Alkali stress may strongly affect assimilation and/or uptake of nitrate. Interference between alkali stress and nitrogen nutrition is a very complex network affecting almost all processes in plant metabolism and development. Thus, nitrogen metabolism regulation may be important in alkali-tolerance as in salt-tolerance. In present study, we chosen rice （Oryza sativa） as the test organism to test the role of nitrogen metabolism regulation in alkali tolerance of rice plants. This article included three experiments:1. a japonica rice variety ’Nipponbare’ was chosen as test organism, and compared effects of salt and alkali stresses on nitrogen metabolism of rice plants;2. using two rice cultivars differing in alkali resistance, we investigated the physiological and molecular responses of nitrogen metabolism to alkali stress;3. the aim of this study was to investigate whether the alkali stress has different effects on nitrogen metabolism in old and young leaves of rice plants, and to compare functions of both organs in alkali tolerance. Major conclusions were as follows:1. Effects of alkali stress on the nitrogen metabolism of ricemainly comprised two mechanisms:①Root. AS caused the reduction of NO3-content in roots, and which caused two baneful consequence, the large downregulation of OsNR1and the subsequent reduction of NH4+production in roots. On the other hand, under AS （pH,9.11）, almost all NH4+were changed to NH3, which caused the severe deficiency of NH4+surrounding the roots. Both events caused the severe deficiency of NH4+in roots. Rice variety ’Nipponbare’ might enhance the expression of OsNRT1;2and OsNRT2;1in roots to increase the frequency of NO3-uptake and to compensate the less of NO3-uptake chemical energy （transmembrane proton gradient）. Under AS, the increased expression of several OsAMT family members in roots might be an adaptative response to the reduction of NH4+in roots or the NH4+deficiency in rhizosphere. Also, the downregulated expression of OsNADH-GOGAT and OsGSl;2in roots might be due to NH4+deficiency in roots.②Shoot. AS caused the large acuumulatiuon of Na+in shoots to toxic level, which possibly affected photorespiration and led a continuous decrease of NH4+production, and inhibited the expression of OsFd-GOGAT and OsGS2in chloroplast.2. Under alkali stress, alkali-tolerant rice was able to maintain normal nitrogen metabolism processes, which might be important for resisting alkali stress.Alkali stress had only small effects on the expression of OsGS2and OsFd-GOGAT in theshoots of the tolerant cultivar, but had clearly reduced expression in the shoots of the sensitivecultivar. Concurrently, alkali stress strongly stimulated the expression of OsGDH1, OsGDH2and OsGDH3in shoots of the sensitive cultivar, but did not affect their expression in shootsof the tolerant cultivar. The above data showed that alkali stress may have changed the NH₄⁺assimilation pathway in shoots of the sensitive cultivar, weakened the frequency of NH₄⁺assimilation by the GS2/GOGAT pathway, and elevated the frequency of NH+4assimilation bythe GDH pathway. Under alkali stress, the down-regulation of OsGS2and OsFd-GOGAT inshoots of the sensitive cultivar might be due to the destruction of the photosynthetic system.Under alkali stress, in shoots, alkali-sensitive rice might accumulate Na+to toxic levels,possibly harming chloroplasts and disturbing relevant metabolism, and immediately reducingPN. The Na+excess in shoots also might influence photorespiration of the sensitive cultivar,and reduce NH+4production from photorespiration, which might immediately down-regulateOsGS2and OsFd-GOGAT in its shoots. Therefore, we propose that down-regulation ofOsGS2and OsFd-GOGAT might be a harmful response of the sensitive cultivar to Na+excessin shoots caused by alkali stress. Na+excess in shoots might even change the pathway ofNH+4assimilation in the sensitive cultivar, weaken the GOGAT/GS pathway and elevate theGDH pathway in roots and shoots. Under alkali stress, alkali-tolerant rice was able tomaintain normal N metabolism processes and relatively high expression levels of OsGS2andOsFd-GOGAT in the shoots, which might be important for synthesis of the N-containingcompounds involved in alkali tolerance.3. Effect of alkali stress on nitrogen metabolism of young leaves was stronger than thatof old leaves.The results showed that alkali stress only produced a small effect on the growth of youngleaves, whereas strongly damaged old leaves. Rice protected young leaves from ion harm viathe large accumulation of Na+and Cl-in old leaves. Compared both tissues, we found that theeffect of alkali stress on nitrogen metabolism of young leaves was stronger than that of oldleaves. Alkali stress did not influence the expression of OsFd-GOGAT and OsGS2in oldleaves, and mightily reduced their expression in young leaves. The decreased expression ofOsFd-GOGAT and OsGS2in young leaves might be a response to NO-3deficiency. The NO-3deficiency in young leaves might cause the large reduction in OsNR1expression and thesubsequent lacking of free NH+4, which might be main reason why alkali stress sharplydownregulated the expression of OsFd-GOGAT and OsGS2in young leaves.