Metabolomic And Transcriptomic Approach To Understanding The Regulation Of Biosynthesis Of Flower Scent In Rose

Flower scent not only improves the ornamental value of the rose but provides aroma materials to perfume and perfume industries.However,most of modern rose varieties lost their flower scent.Studies have shown that the transcriptional regulation can change flower scent composition efficiently,and therefore is promising to break through the shortcomings and limitations from traditional cross breedings.But few studies are available on transcriptional regulation of flower scent biosynthesis at present.In this study,a new rose cultivar’Tianmidemeng’,which was developed from the male parent ’Elle’ and female parent ’First blush’by our group,together with the parent cultivars and the other F1 offspring ’Qingge’,were used to explore the molecular basis of flower scent heterosis in ’Tianmidemeng’.The main results are as follows:1)In order to grasp the key timepoint or period of flower scent synthesis accurately,we studied the variation of flower scent emissions following the daily rhythm and flowering process with ’Tianmidemeng’ as material.The results showed that the flower scent emission presented obviously diurnal and developmental changes,in which it reached the highest at 12:00 and at semi-flowering stage,respectively.2)Compared to parents,synthesis levels of terpenoids,especially monoterpene alcohols and their derivatives have been significantly improved in ’Tianmidemeng’,which contributed to the formation of flower scent heterosis.Many more non-additive genes,especially the up-regulated genes were expressed in ’Tianmidemeng’ than ’Qingge’,of which GO functional enrichment showed that the‘terpenoids biosynthesis’,’transport,signal transduction and response pathways of auxin’,and ’transcription activity’ were changed significantly in ’Tianmidemeng’.Expression changes of genes involed in flower scent biosynthesis and auxin-production/response pathways supported the results.Differentially expressed genes(DEGs)between parents were enriched in ’Tianmidemeng’,of which the number is much larger than ’Qingge’;however,in the DEGs the parental expression level dominance(ELD)genes might not cause the formation of flower scent heterosis directly,but contributed to a compromise between the improved flower scent and reduced resistance;transgressive genes promoted metabolic pathway changes in’Tianmidemeng’,possibly via a way mediated by auxin response pathway and transcriptional activity.Significant difference was found in transcription factors(TFs)between parents,while these differentially-expressed TFs were enriched in "Tianmidemeng",supporting the deduction that TFs may play important role in flower scent heterosis.3)Using weighted gene co-expression network analysis(WGCNA)method,DEGs among four cultivars and three flowering stages of’Tianmidemeng’,including TFs,were correlated to 19 flower scent components from four cultivars,aiming to identify candidate genes(TFs)related to monoterpene alcohols,phenylpropanoid/benzenoids.Important modules from the analysis,such as white and midnightblue modules,both have enriched terpenoids metabolic pathways and auxin-responsive pathways,among which the TFs were important candidates for further research.4)A novel "grafting VIGS" system was set up with the help of grafting in this study,and was used to successfully silence the geraniol synthase gene RhNUDX1 in Rosa chinensis ’Old blush’ flowers.With this method,we can observe phenotypes of silenced flowers within 4 or 5 weeks,60-240d earlier than tranditional method.In addition,the silencing effenciency of 47.4%was greatly improved compared to that of 10%-35%with tranditional method.5)RhNUDX1 promoters were cloned from ’Tianmidemeng’,the parents and ’Qingge’,respectively.No distinctive changes were generated in RhNUDX1 promoter of ’Tianmidemeng’;but binding elements of MYB TFs and auxin response elements were involved in it,supporting the above inference for the heterosis formation.Seven TF genes predicted by WGCNA were cloned and overexpressed in tobacco leaves.It was found that 1 AP2/ERF and 1 MYB TF genes,TmMYBl and TmERF1 increased emission leves of geraniol isovalerate in tobacco leaves,indicating that terpenoids synthesis might be regulated.By the "grafting VIGS",the silenced flowers of TmMYBl gene produced much less geraniol compared to control flowers,further surporting its role in regulation of terpenoids production.However,the 2 TFs do not directly bind to the promoter of RhNUDX1.To sum up,the transgressive genes,ELD genes and non-additive genes may contribute to the formation of flower scent heterosis in ’Tianmidemeng’ together,and the mechanism may be related to the alterations of auxin-response pathways and transcription factors.Based on it,the WGCNA was used to identify TFs involved in flower scent synthesis.The experiment results verified the accuracy of the prediction,and 2 TF genes were identified.This study provides an important reference for the further research to regulation of flower scent synthesis in rose.Meanwhile,it provides ideas and candidate genes for flower scent improvement by transgenetic approach in rose.