Preliminary Study Of Proteomics Of Pyropia Haitanensis In Response To Desiccation Stress

Pyropia haitanensis is one of the most economically important mariculture crops of southern China,accounting for 75%of the total output of cultivated Pyropia in China.P.haitanensis is also considered to be a research model for studying physiological ecology and the molecular mechanisms of stress tolerance of intertida seaweed communities.To understand the important biological characteristics of P.haitanensis,some related researches,such as EST and transcriptome sequencing,mitochondrialand chloroplast genome sequencing,have been completed.Transcriptome and expression profile analysis in response to desiccation stress in P.haitanensis have also been characterized.Nevertheless,research on P.haitanensis still relatively limited.In this study,the differential protein expression of P.haitanensis（Z-61）blades under different dehydration level（0,10%,40%,60%,90%）stress and rewatering were studied.At the same time,the changes of some physiological indices of antioxidant system and photosynthetic system at the same level stress have also been determined.The main results are as follows:（1）The results of physiological indices under different levels of dehydration stress showed that 60%of the water loss of P.haitanensis blades was an important boundary.When the water loss<60%,the major constituent of produced ROS in P.haitanensis cell was H₂0₂,which is the low toxicity ROS,and the contents of high toxicity ROS,O₂^-·,has no significant changes.For the constituent of antioxidant system,the activity of APX and contents of ASA and GSH were no significant changes,so much as the activities of SOD,CAT and GR were significant down-regulated.But when the water loss>60%,as the down-regulation of H₂0₂ content,the content of high toxicity O₂^-·was significant up-regulated and became the major constituent of ROS in P.haitanensis cell,although the activities of SOD,CAT and APX still were no significant up-regulated,the activity of GR and contents of ASA and GSH were significant up-regulated.These results suggest that GR,ASA and GSH play important roles in cleaning the ROS during serious desiccation stress and the antioxidant enzymes of SOD,CAT and APX have no function in ROS cleaning during desiccation stress.（2）The results of chlorophyll fluorescence parameters under different levels of dehydration stress showed that:qN is significantly decreased under mild dehydration（water loss rate<35%）,ETR is significantly decreased under moderate dehydration（water loss rate was 40⁶0%）stage,and Fv/Fm,Y（II）,qP is significantly decreased under severe dehydration（water loss rate>60%）.qN reached its lowest value under 70%water loss rate,ETR,Y（II）,qP reached their lowest point under 80%water loss rate;and Fv/Fm still decreased significantly during water loss rate more than 90%.These results indicated that P.haitanensis could maintain the normal operation of photosynthesis by adjusting photosynthetic organ function and reducing heat dissipation under mild and moderate water loss stress,and during severe water loss stress,the PSII system,photosynthetic organ and photosynthetic enzyme system of P.haitanensis were damage,but these damages were reversible,their activity could be fully restored when re-submerged.（3）Compared the 2-DE map of Z-61 blades under different levels of dehydration（0,10%,40%,60%,90%）stress and rewatering,a total of 100 protein spots were selected for their expressed level changing（up-regulation or down-regulation）more than twice.By analyzing with mass spectrometry and blasting with the public database and transcriptome database of P.haitanensis,68 differentially expressed proteins were identified successfully.These differentially expressed proteins were related to photosynthesis and energy metabolism,nitrogen,amino acids and protein metabolism,ascorbate and aldarate metabolism,biosynthesis of terpenoids and steroids,cytoskeleton,transcription and translation,chaperone protein and other life processes.The different expression of key proteins were further verified by western blot.（4）The results of physiological indices,chlorophyll fluorescence parameters and different proteomics suggested that 60%of the water loss of P.haitanensis blades was an important boundary.When the water loss<60%,the photosynthetic rate,antioxidant enzyme system and material metabolism have no significant change,but when the water loss>60%,the photosynthetic rate and material metabolism were significant down-regulated,and the activity of antioxidant enzymes was significant up-regulated.