Research On Cell Protection Mechanism During Maturation Drying In Nelumbo Nucifera Embyro

When the lotus（Nelumbo nucifera）embryo matures,it loses about 90%of its moisture and could maintain cell viability for a long time.It is the longest life form that has been scientifically determined.The storage tolerance of lotus embryo is closely related to its resistance to stress during maturation drying.In this study,three aspects of morphological observation,physiological index determination and proteomics analysis were used to study the cell protection mechanism of lotus embryo during mature drying.The morphological study showed that the lotus seed embryo reached morphological maturity on the 15th day after pollination;the embryo fresh weight reached the maximum on the 21st day after pollination,and its dry weight had reached 73.3%of the mature embryo,which means the nutrients of lotus embryo are in a vigorous accumulation and gradually enter the stage of maturation drying.From 21 to 40 days after pollination,the moisture content of lotus embryo plummeted from 73.7%to 10.2%,accompanied by continuous accumulation of soluble sugar,protein and proline,the seed embryo reached fully maturity,and entered the physiological dormant period.The physiological measurement of cell MDA,ROS components and antioxidant system showed that the oxidative stress of lotus embryo during continuous dehydration increased significantly,the most severe stress happened on the 27th day after pollination and decreased subsequently;correlation analysis results of various physiological indicators of resistance showed that H₂O₂ is the main ROS component during the maturation drying of lotus embryos,and a large amount of proline and antioxidants such as GSH,V_E accumulated in the lotus seed embryos during maturation drying played an important protective effect on cells damaged by stress.A total of 815 differentially expressed proteins（DEPs）were selected from the three stages of lotus embryo by tandem mass tags（TMT）technology.Compared with 21 days after pollination（DAP）,27 DAP and40 DAP have 756（up 198;down 558）and 618（up 198;down 420）DEPs,respectively.GO functional annotation indicates that up-regulated DEPs in drying seed embryo are mainly involved in response to stimulus stress,and down-regulated DEPs are mainly involved in cell metabolism and synthesis;KEGG enrichment pathway analysis indicates that sugar metabolism,protein processing and synthesis,glutathione metabolism activities are enhanced in lotus embryo during maturation,while cell metabolism,photosynthesis and fatty acid metabolism pathways are significantly reduced;protein interaction network analysis shows that lotus embryo are mainly involved in activities such as redox homeostasis,carbohydrate metabolism,fatty acid metabolism and cell wall formation,in addition,metabolic process is a specific biological process in seed embryo cells during the rapid stage of dehydration,response to ABA,protein modification and processing are specific processes in seed embryo cells during the late stage of dehydration.According to the results of bioinformatics analysis,7 biological processes that are closely related to the stress resistance of lotus embryo during maturation drying were found,on this basis,145 key functional proteins were further selected,which were involved in carbohydrate metabolism（36）,fatty acid metabolism（17）,redox homeostasis（15）,stress defense processes（35）,protein modification and degradation（13）,hormone response（7）,and DNA/RNA repair（22）.The parallel reaction monitoring（PRM）results showed that the protein expression level was basically consistent with the TMT identification results,suggesting that the TMT proteomics identification results were credible for further analysis of DEPs.This study proves that there are more complex molecular regulatory network mechanisms of lotus embryo cells during maturation drying.The cell mainly provides energy for various substances through glycolysis,pentose phosphate pathway and galactose metabolism activities related to raffinose oligosaccharide family synthesis,and improves cell resistance to stress during continuous dehydration;antioxidant system represented by TRX and GSH,a large number of DEPs related to protein modification and degradation,stress defense,DNA and RNA repair play an important protective role in dehydrated lotus embryo cells;in addition,the development of lotus seed embryo cells may be regulated by the ABA signaling pathway.The energy metabolism of various substances in cells and the defense and protection system work together to improve the cell resistance and storage tolerance of dehydrated lotus embryos.