The Response Of The Key Photosynthetic Enzymes In Caragana Korshinskii Under Natural Increased Drought Stress

Drought is one of the most important environmental factors that affects the survival, growth and development of plants. In order to adapt to water deficit, plants that grown under drought conditions need certain morphological, physiological and molecular mechanisms to hold enough water to maintain normal physiological metabolism. Researches about the mechanism of drought stress led to a series of complex changes in plant physiology from the molecular to the ecological focused on aspects of morphology and anatomy in particular drought physiology, as well as geographic distribution and ecological characteristics, but researches of the molecular mechanism of the formation of plant resistance can not explain the drought resisting of xeric plants fundamentally. To elaborate the mechanism of plant drought resistance, a new view has been proposed that plants in order to achieve the adaptation of arid conditions to survive and growth will shift their photosynthetic carbon assimilation pathway to C4 pathway. Currently, researches about plants shift to C4 pathway to resistant drought stress include morphology changes of assimilation organ, changes of photosynthetic enzyme activity and changes of stable carbon isotope. However, an activity of the C4 photosynthetic key enzymes is crucial to improve the development of C4 pathway.In order to clear the physiological and biochemical changes occurring in the photosynthetic enzymes and clarify the appropriate drought mechanism of development of C4 pathway of xerophytes, we have chosen Caragana korshinskii that widely distribute in the Loess Plateau and acte as an advantage farm Land vegetation legume genus as our research material, and use the natural formation of the drought gradient as a condition of processing. In order to provide experimental supports of the theory that C3 plants will shift to C4 photosynthetic pathway to increase the drought resistance, we have explored the change of the distribution, content and expression of photosynthetic key enzymes of Caragana korshinskii by molecular biology, plant physiology, morphology and anatomy methods.On the basis of transcriptome prediction studies of Caragana korshinskii blade, the present experimental results showed that:(1) The results of gene cloning and alignment analysis of sequences of photosynthetic critical enzymes showed that the similarity of cloned pepc sequence with proved C4-type gene sequence is as high as 90%, and pepc gene has begun to have a molecular characterization of C4-type pepc genes; phylogenetic tree based on entire amino acid sequence of the PEPC and the large subunit of Rubisco also showed Caragana korshinskii acts between C3 plants and C4 plants, and keep a short distance with drought resistance C4 woody plants. The results indicated that Caragana korshinskii already have developed molecular basis of photosynthetic enzymes to develop the C4 pathway.(2) Quantitative PCR results showed that the expression level of photosynthetic key enzyme genes pepc, nad-me and ppdk act an increasing trend with increased drought stress; but rbc L and pepck were relatively stable at the level of transcription; and nadp-me shows a downward trend. Western blotting results showed that protein expression level of photosynthetic key enzyme PEPC, NAD-ME, PPDK act gradually increased with increased drought; but the expression of NADP-ME showed decreased; the expression of Rbc L and PEPCK has not a significant change. Enzyme activity measurement and analysis showed that the activity of photosynthetic key enzyme PEPC, NAD-ME, PPDK increased with increased drought stress; but the activity of NADP-ME and PEPCK is reduced with increased drought stress; and the activity of Rbc L acted elevated at first and then decreased. The relative content of protein and activity analysis showed that the C4 pathway that in the development of Caragana korshinskii is NAD-ME subtype.(3) The result of anatomical structures showed that the size of bundle sheath cells increased under drought conditions, indicating Caragana korshinskii has begun forming morphological characteristics that to adapt to the C4 pathway; the analysis of stable carbon isotope of Caragana korshinskii blade in different rainfall areas showed that the value of δ13C that can indicate a long-term water use efficiency increase with increased water deficit conditions, this result will give a physiology evidence of Caragana korshinskii to develop the C4 pathway.Based on above results we can obtain the following conclusions: an important part of the mechanism of adaptation to drought of Caragana korshinskii is the positive response of C4 photosynthesis critical enzymes in transcription and translation level under naturally drought conditions. Caragana korshinskii obtained higher net photosynthetic rate and accumulate more biomass to improve the survival and growth capacity through the development of the C4 pathway, this conclusion is also consistent with previous studies in our laboratory.