| Phenolic compounds are one of the most important secondary metabolites which have important physiological functions in diverse aspects of grape berry development,such as pigmentation and co-pigmentation,free radical scavenging,ultraviolet(UV)radiation protection and defense against pathogen infections.Furthermore,their contribution to the color,bitterness,astringency and antioxidant properties of red wine and potential benefits for human health have gained much attention on studying the biosynthesis of phenolics in grape berries over the years.However,the regulatory mechanism of berry phenolic metabolism under different environment conditions remains largely unstudied.In the present study,we constructed a robust reference transcriptome with a non-redundant set of 52,359 transcripts for berries of Cabernet Sauvignon and provided the transcriptome comparation at different developmental stages of the berries from eastern and western China through next-generation sequencing technology.The regional differentially expressed genes from grape berries were significantly enriched in the phenolic biosynthetic pathway at each developmental stage,which might be caused by the environmental deviations such as light condition,temperature and water status between the two regions.Chosen light factor as research emphasis,we subsequently analyzed the accumulation and compositional changes of hydroxycinnamic acids(HCAs)and flavonoids,together with the expression profiles of phenolic biosynthetic related structural and regulatory genes in grape berries under various cluster sunlight exposure and exclusion treatments at the metabolomic level and the transcriptomic level,respectively.The results exhibited that the expression level of genes encoding trans-cinnamate 4-monooxygenase(C4H),4-coumarate:CoA ligase(4CL)and cinnamoyl-CoA reductase(CCR)family members and concentration of HCAs were significantly increased in the sunlight exposure berries in comparison with the control berries at both veraison and harvest,while sunlight exposure and exclusion treatments obviously stimulated and inhibited the expression of genes encoding flavonol synthase(FLS)family members and the accumulation of flavonols in berry skins.Further coexpression analyses indicated that some members of MYB,bHLH,NAC,ERF and C2H2 transcription factor families may positively or negatively regulate both the flavonol and HCA biosynthetic pathways in grape berries in response to light.Based on the grape reference genome and transcriptome sequences,a bioinformatics analysis of the systematic evolution and gene structure of phenylalanine ammonia-lyase(PAL)family in the Cabernet Sauvignon grape was conducted in detail.Expression profiling reveals that VviPAL2 and VviPAL15 were strongly expressed in various grape tissues,and their expression throughout berry development was positively correlated with the accumulation of phenolic compounds.Besides,changes in transcript abundance of VviPAL2 in berries under sunlight treatments and VviPAL15 in berries under temperature stress were also correlated with the biosynthesis of phenolics in grape berries.In addition,the gene structure,expressional characterization of VviFLS4 and VviFLS5,as well as the enzyme activity of their coded proteins were analyzed systematically.Several putative transcription regulators of VviFLS4,including VviERF5,VviHAT5,VviMYCA1 and VviMYBF1,were also screened and validated through yeast one-hybrid and vivo/vitro experiments.Taken together,our study provided a valuable overview of the phenolic metabolism and transcriptome changes under different ecological conditions and light treatments,and also characterized the biological functions and transcriptional regulations of grape PALs and FLSs,which provide new valuable insights into the regulation of flavonoid metabolism in grape berries. |
