Proteomic Investigation Of Peach Fruit During Ripening And After NO Treatment

Peach(Prunus persica L.),with economic value,is the most important stone fruit in the world and in China.High-quality honey peach fruit in Jiangsu and Zhejiang all has short shelf-life at ambient temperature,which causes the product to be restricted in the market circulation and short the supply period.In recent years,the research on the mechanism and technology of fruit ripening and senescence has attracted much attention.This thesis is about the identification and functional analysis of differently expressed proteins during peach fruit ripening as well as their regulation by NO treatments,in order to further explain the molecular mechanism of the biological process of peach fruit ripening and the effect of NO treatment,and also to provide a reference for the comprehensive and in-depth study on the molecular mechanism of ripening of peach fruit.The main results are as follows:1.'XiaHui No.8' peach fruit in different harvest and riping time were analyzed by iTRAQ proteomic technology.There are 5047 proteins were found and 1207 proteins of them were identified as three biological repeated samples in common.There are 387 protein abundance changes more than 1.2 times(including 79 differentially expressed proteins for more than 2-fold and 62 differentially expressed proteins for more than 1.5 times or 2 times).These protein belong to 21 functional categories,which are mainly involved in the Posttranslational modification,protein turnover and chaperones(12.81%)and Carbohydrate transport and metabolism(11.70%),Amino acid transport and metabolism(7.80%)\Translation,ribosomal structure and biosynthesis(6.13%)and Energy production and conversion(6.13%).2.According to the analysis of peach protein in different harvest and riping time,in group C/A?C/B?D/C?E/C and F/C,there were 23,1,23,42 and 3 difference proteins abundance more than 2-fold,respectively.Among them,the C/A and E/C groups have the most functional categories.C/A group 1.2-fold differential protein number is five times more than C/B group(165/32),2-fold differential protein number is 23 times more than C/B group(23/1);E/C group 1.2-fold differential protein number is 2.90 times(203/70)and 5.34 times(203/38)more than group D/C and F/C,2-fold differential protein number is 1.83 times(42/23)and 14 times(42/3)more than group D/C and F/C,respectively.These results showed that there were significant differences in the biological pathway,orientation,intensity and change of peach fruit ripin.3.According to the analysis of protein function,pre-harvest peach fruit protein function mainly related to synthesis of sugars and carbohydrates,protein translation,expression,and modification,with strong capability of ion and material transportation,which effectively maintain osmotic equilibrium,and also with strong oxidation reduction reaction.The Rubisco in on-harvest(Close to harvest)peach fruit begins to degrade to provid nitrogen source for the synthetic reaction.The peach fruit(in the harvest time)oxidative phosphorylation and respiration have increased.Metabolism and synthesis of amino acid have exacerbate.Sugar and carbohydrate metabolism were mainly of decomposition.Post-riping peach fruit have very strong oxidation reduction reaction ability and the reaction of oxidative phosphorylation reach a peak,have strong ability of material transportation and lipid metabolism and protein translation activityare extremely active,cell wall degradation rate accelerated.4.The extracted proteins from the NO peach fruit(10 ?L/L 3h and then stored at room temperature 20-25 ?,'Xiahui No.5' peach fruit)and CK were used for proteomic analysis in order to understand the response of post-harvest peach fruit to NO treatment during ripening stage at proteomic level.After two dimensional gels electrophoresis(2-DE)was conducted,differently expressed spots(P<0.05)were selected to be excised and analyzed using MALDI-TOF/TOF,and finally 104 protein spots were confidently identified according to NCBI database.The results demonstrated that 104 protein spots are involved in different metabotic process,which can be classified into seven classes according to their functions,among which 30.77%were related to energy and metabolic,25.00%were related to stress response,8.65%to cell structure,8.65%to protein fate,6.73%to transportation and transduction and 5.77%to ripening and senescence etc.In differential protein,metabolism of energy and protein proportion has the largest rate,and the second is involved in energy production,material(carbohydrates,amino acids,protein,lipid)and secondary metabolism.These results suggest that NO deal with significant impact on the protein expression of energy and metabolism and related stress reaction and the defense related,and then participate in the process of peach fruit riping.5.Cell location results showed that most of the protein is located in cytoplasm(40)and chloroplast(34),the nucleus(10),mitochondria(8)and extracellular space(3),vacuole(2),the colored body(1)and endoplasmic reticulum chamber(1),which indicates that the effect of NO treatment on peach protein expression is associated with many organelles.6.According to the analysis of protein function,,the regulating mechanism of NO on peach fruit is complex,NO could largely up-regulate the abundances of SOD and HSP70 to effectively induce the defense capacity through promoting the production of the complex ACO-NO-ACC to affect ethylene biosynthesis,depressing the synthesis of cytochrome c oxidase to inhibit electron transport and oxygen consumption,and strengthening the structural components "profilins" to repress the degradation of cell wall.In addition,NO could induce the abundance increase of SOD and the ascorbate-glutathione cycle-involving enzymes to inhibit the generation and accumulation of ROS,and repress the degradation of cell wall through depressing the loss of Ca2+ ions and other structural components to maintain the mechanical properties of cell structure and then cell's function.