Metabolic Response At High Temperature And Activity Of Leaf Constituents Of Five Raspberry Cultivars

Raspberry(Rubus idaeus L.)is known as one of the most famous healthy fruits and is consumed fresh or processed product all over the world.Raspberries are suited to warm and humid environments,and their low tolerance to high temperatures has become one of the most important factors limiting the widespread cultivation of raspberries.The flowering period is the period when raspberries are very sensitive to temperature changes,and is also the most critical stage to determine the yield of raspberries.Therefore,it is of great significance to explore the key metabolites and gene resources of heat resistance of raspberries through the study on the metabolic response mechanism of high temperature in the flowering period for the breeding and planting industry development of raspberries.Second,raspberry leaves are rich in active metabolites,and the comparison of the content and types of metabolites in different raspberry cultivars leaves will help to utilize the resources raspberry leaves.In this study,the leaves of five different raspberry cultivars,‘Autumn Britten’(ABr),‘Autumn Bliss’(ABl),‘Red Autumn’(RA),‘Heritage ’(HR),and ‘Korean Black’(KB)were used as materials to investigate the resistance characteristics of the leaves under high temperature stress and the antioxidant activity of the active extracts using metabolome,transcriptome and functional validation of the active extracts.The main results are as follows:1.The metabolite content of different raspberry cultivars changed after high temperature stress and there were significant differences,among species.213 metabolites were analyzed for significant differences in sugars,lipids,and amino acids,and the metabolic responses of ‘Autumn Bliss’ and ‘Korean Black’ cultivars to high temperature stress were higher than those of other cultivars.2.A total of 748 significantly different metabolites between species were determined from the metabolome analysis of normally growing raspberry leaves of different species,with lipids,amino acids and their derivatives,sugars,and flavonoids accounting for the largest proportion of different metabolites.3.Transcriptome analysis of different leaves of raspberry cultivars after high temperature stress revealed the screening of 1332 up-regulated differential genes enriched in a total of 125 metabolic pathways and 720 downregulated differential genes enriched in a total of 121 metabolic pathways.Metabolic pathway enrichment results mainly included amino acid metabolism,sugar metabolism,lipid metabolism,energy metabolism,and the secondary metabolite synthesis.Transcription factor analysis identified four genes belonging to the heat-stimulated transcription factor(HSF)family in each of the‘ Autumn Bliss’ and ‘Korean Black’.The most differential genes were found in ‘Korean Black’ and the fewest in ‘Heritage’.Pathway enrichment analysis of the differential genes specific to these two cultivars revealed that both responded to the α-linolenic acid metabolic pathway after heat stress.4.The combined metabolome and transcriptome analyzes revealed that the major metabolic pathway for raspberry leaf to respond to high temperature stress is the regulation of the metabolite content of amino acids,sugars,and lipids by transcription factor family genes such as WRKY,AP2/ERF-ERF,MYB,NAC,b HLH and HSF,as well as metabolic pathways such as amino acid metabolism,glucose metabolism,lipid metabolism and energy metabolism to adapt to high temperature stress.Analysis of specific differential metabolites and differential genes ‘Korean Black’ and ‘Heritage’and the q RT-PCR validation of key genes expression showed that differential expression of genes and metabolites related to the α-linolenic acid metabolic pathway between cultivars may be key to influencing the response of raspberry to high temperature stress.5.Quantitative comparison of Ellagic acid,Raspberry ketone,Epicatechin and Asiatic acid in raspberry leaves after high temperature stress and during different growth periods showed that high temperature stress and time accumulation significantly increased the content of all four active components in most cultivars.6.Alcoholic extracts of raspberry leaves of different cultivars from September were selected to compare the levels of superoxide dismutase(SOD),glutathione(GSH),catalase(CAT),and reactive oxygen species(ROS)before and after pre-protection of L929 fibroblasts by leaf extracts.This confirmed that alcoholic extracts of ‘Korean Black’ raspberry leaves were effective in protecting L929 fibroblasts from UVB-induced damage.