Transcriptomic And Proteomic Analysis Of Golden Queen Hami Melon Under Cold Stress

Hami melon（Cucumis melo var.saccharinus）is favored by consumers because of its sweetness and flavor.As a characteristic fruit of Xinjiang,China,Hami melon plays a significant role in promoting the advancement of the local economy.After postharvest,Hami melon is stored at low-temperature for preserving its commercial quality.Long terms storage,however,gives rise to cold injury including browning,pitting,etc.,adversely affecting the storage quality of Hami melon.Therefore,studying the molecular mechanism of Hami melon response to cold stress has important theoretical and practical significance for maintaining the quality of Hami melon and prolonging the shelf life.In this thesis,the representative local variety"Golden Queen"was used as the test material to observe and analyze the phenotype,physiological and biochemical indexes,and studied the occurrence of chilling injury and the changes of related indexes stored at 0.5℃,3℃and 21℃（control）.Then,the differentially expressed genes（DEGs）and differentially expressed proteins（DEPs）of Golden Queen Hami melon under different storage temperatures were found by transcriptome and proteome sequencing.The important cold-resistance genes MYB and heat shock proteins were screened and mined,and their functions in Hami melon in response to cold stress were analyzed.This study provides a theoretic basis for further investigating the molecular mechanism of cold resistance in post-harvest Hami melons,and has broad implications for food science and plant breeding.The specific results are as follows:（1）Phenotype features showed that compared with Hami melon stored at 0.5 and 21℃,Hami melon at3℃processing a higher storage quality,which indicated that the cold injury was correlated with storage temperature and time.Though both Hami melon stored at 0.5 and 3℃suffered a cold injury,the symptoms of cold injury at later one presented at a late period with lower rating rate,weight loss,cold injury index.Lower H₂O₂ content,MDA content,lipoxygenase（LOX）activities,and higher peroxidase（POD）,catalase,（CAT）superoxide（SOD）,fatty acid desaturase（FAD）and adenosine triphosphatase（ATPase）activities,resulting in the alleviation of cold stress and maintaining the quality of Hami melon.（2）Transcriptomics analysis indicated that 6479（up 1314,down 5165）,7914（up 1594,down 6320）,7350（up 1874,down 5476）,6482（up 2751,down 3731）,3642（up 1638,down 2004）and 3532（up 1198,down 2334）DEGs were identified at 0.5℃/12d-0d,0.5℃/24d-0d,3℃/12d-0d,3℃/24d-0d,21℃/12d-0d,21℃/24d-0d,respectively.A total of 415 DEGs were conmen expressed at above six groups.Gene Ontology（GO）annotation analysis showed that the DEGs were mainly annotated to‘metabolic process’,‘cellular process’,‘response to stimulus’,‘cell’,‘cell part’,‘membrane’,‘catalytic activity’and‘binding’.The kyoto encyclopedia of genes and genomes（KEGG）metabolic pathway showed that the main metabolic pathways involved in the DEGs were:‘metabolic process’（ko01100）,‘plant-pathogen interaction’（ko04626）,‘starch and sucrose metabolism’（ko00500）,‘plant hormone signal transduction’（ko04075）,‘fatty acid metabolism’（ko01212）and‘biosynthesis of secondary metabolites’（ko01110）,etc.（3）A total of 1163 differentially expressed transcript factors including MYB（375,32.24%）,AP2/EREBP（86,7.39%）,b HLH（63,5.42%）,FAR1（56,4.82%）,NAC（56,4.82%）and WRKY（50,4.30%）were identified.These differentially expressed transcription factors were mainly involved in the process of transcriptional regulation,response to stress and metabolism.（4）Based on the RNA-seq analysis,28 MYB genes were identified.Two stress-related MYB genes,JhMYB24 and JhMYB48 genes were analyzed.The results showed that the proteins encoded by the JhMYB24and JhMYB48 genes belonged to the R2R3-MYB type transcription factors,and their amino acid sequences possessing typical R2R3-MYB structure were highly conserved with the Arabidopsis thaliana,melon and summer squash MYB,reveling the similar functions in the process of resisting stress.The results of q RT-PCR showed that the expression levels of JhMYB24 and JhMYB48 were consistent with the transcriptome sequencing results.（5）Proteomic analysis showed that 360（down 218,up 142）,447（down 287,up 160）,600（down 250,up 350）and 638（down 277,up 361）,598（down 199,up 399）and 950（down 458,up 492）DEPs were isolated at 0.5℃/12d-0d,0.5℃/24d-0d,3℃/12d-0d,3℃/24d-0d,21℃/12d-0d,21℃/24d-0d.The results of GO analysis showed that the DEPs were mainly annotated to‘metabolic process’,‘response to stimulus’,‘cell’,‘catalytic activity’and‘binding’.COG analysis found that DEPs were mainly related to‘posttranslational modification’,‘protein turnover’,‘molecular chaperones’,‘translation’,‘carbohydrate transport’and‘metabolism’.KEGG metabolic pathway enrichment analysis showed that DEPs were enriched in‘carbon metabolism’（ko01200）,‘metabolic pathways’（ko01100）,‘phenylpropane biosynthesis’（ko00940）,‘glutathione metabolism’（ko00480）,etc.A total of 81 DEPs related to stress like 38 heat shock proteins（HSPs）,five peroxidases,four glutathione S-transferases（GSTs）were obtained.（6）Based on the results of i TRAQ sequencing（proteomic）analysis,38 HSPs in response to stress were identified.The alignment analysis of four HSPs（JhsHSP,JhHSP70,JhHSP90Ⅰand JhHSP90Ⅱ）showed that they were highly conserved with HSPs in cucumber,cocoa,balsam pear and Cucurbita pepo.The promoters of JhsHSP,JhHSP70,JhHSP90Ⅰand JhHSP90Ⅱall contained low-temperature stress response elements（LTRE）,indicating that these proteins played an active role in the response to cold stress of Golden Queen Hami melon.The results of q RT-PCR were consistent with the results of proteomics.The expression levels of JhHSP70 and JhHSP90I in the Golden Queen Hami melon stored at 3°C were much higher than those at0.5°C and 21°C,indicating that they may play an important role in the response to cold stress.