Physiological Response Of Highland Barley To Low Nitrogen Stress And Studies On The Mechanism Of Alternative Respiratory Pathway Affecting Low Nitrogen Tolerance

Low nitrogen(N) stress is a major limiting factor for plant growth and crop productivity. Studying on the mechanisms of crop tolerance to low-N stress can provide a theoretical basis for the breeding of new cultivars with high nitrogen use efficiency. Highland barely is widely cultivated in the Tibetan Plateau of northwestern China. Due to the complexity of the plateau climate, highland barley has strong tolerance to environmental stresses. Highland barely is an ideal material to study the mechanisms of crop tolerance to environmental stresses. In this study, we used highland barley(Kunlun12) as the material and barley(Ganpi6) as the control to study the physiological responses to low-N stress and the role of alternative respiratory pathway under low-N stress for understanding the tolerance mechanism of highland barley to low-N stress. The main results were summarized as follows:1. Low-N stress inhibited growth, reduced the contents of nitrate, soluble protein and chlorophyll, and decreased photosynthesis in highland barely. Compared to Ganpi6, Kunlun12 showed a relatively higher growth rate, nitrate, soluble protein and chlorophyll contents, and photosynthesis under low-N stress. These results suggested that Kunlun12 has stronger tolerance to low-N stress than Ganpi6.2. In both Ganpi6 and Kunlun12, low-N stress promoted the growth of primary roots and increased the ratio of root/shoot, which are conducive to absorb more nitrogen. These effects were more obvious in Kunlun12 than in Ganpi6. Under low-N stress, the length of primary roots and the ratio of root/shoot increased by 101.7% and 83.9% in Kunlun12,respectively; they increased by 53.9% and 50.1% in Ganpi6,respectively. This could be one of the reasons why Kunlun12 has stronger tolerance to low-N stress than Ganpi6.3. Nitrate reductase(NR) and glutamine synthetase(GS) are two key enzymes in the process of nitrogen assimilation. The activities of NR and GS decreased under low-N stress in both Ganpi6 and Kunlun12, but the decrease in Ganpi6 was more than that in Kunlun12. Under low-N stress, the activities of NR and GS decreased by 59.9% and 33.8%, respectively, in roots of Ganpi6 and 82.2% and 24.4%, respectively, in leaves of Ganpi6; and they decreased by 31.3% and 18.1%, respectively, in the roots of Kunlun12 and 47.9% and 11.8%, respectively in the leaves of Kunlun12. These results suggested that Kunlun12 has better capacity in nitrogen assimilation under low-N stress, which may be another reason why Kunlun12 has stronger tolerance to low-N stress than Ganpi6.4. Low-N stress led to excessive production of reactive oxygen species(ROS) in both Ganpi6 and Kunlun12,resulting in membrane lipid peroxidation. Under low-N stress, Ganpi6 produced more ROS, resulting in severe membrane damage, which was more obvious in leaves than in roots. Under low-N stress, the activities of antioxidant enzymes and glucose-6-phosphate dehydrogenase(G6PDH) in roots increased in Ganpi6 and Kunlun12 to help scavenge ROS. But the activity of G6 PDH did not increase in leaves, indicating that the function of G6 PDH is different in roots and leaves.5. Under diverse stresses,alternative respiratory pathway is induced, helping control the generation of ROS. Under low-N stress, the alternative respiratory pathway increased by 28.6% in leaves of Ganpi6, but increased by 42.0% in leaves of Kunlun12; the levels of alternative oxidase(AOX) protein increased significantly in leaves of Kunlun12 but not in Ganpi6; the alternative respiratory pathway did not increase in roots. These results indicated that alternative respiratory pathway has different functions in roots and leaves.6. Under low-N stress, NADPH accumulated in chloroplasts of Ganpi6 and Kunlun12, leading to inhibition of photosynthesis and excessive production of ROS. This phenomenon was more obvious in Ganpi6 than in Kunlun12. The excessive NADPH produced in chloroplasts could be transported through Malate-OAA shuttle into cytosol or mitochondria, where it is consumed through alternative respiratory pathway. So alternative respiratory pathway could reduce the accumulation of ROS and protect photosynthesis. Under low-N stress, the alternative respiratory pathway had more increase in leaves of Kunlun12 than in Ganpi6, so it could better dissipate the reducing equivalents from chloroplasts, thereby reducing the accumulation of NADPH in chloroplasts and the generation of ROS. So Kunlun12 has less oxidative damage, which might be another reason for the tolerance of Kunlun 12 to low-N stress.