Using the combined analysis of transcripts and metabolites to propose key genes for differential terpene accumulation across two regions

Ya-Qin Wen, Gan-Yuan Zhong, Yuan Gao, Yi-Bin Lan, Chang-Qing Duan, Qiu-Hong Pan

Terpenes are of great interest to winemakers because of their extremely low perception thresholds and pleasant floral odors. Even for the same variety, terpene profile can be substantially different for grapevine growing environments. Recently a series of genes required for terpene biosynthesis were biochemically characterized in grape berries. However, the genes that dominate the differential terpene accumulation of grape berries between regions have yet to be identified. Free and glycosidically-bound terpenes were identified and quantified using gas chromatography-mass spectrometry (GC-MS) technique. The transcription expression profiling of the genes was obtained by RNA sequencing and part of the results were verified by quantitative real time PCR (QPCR). The gene co-expression networks were constructed with the Cytoscape software v 2.8.2 ( www.cytoscape.org ). 'Muscat Blanc a Petits Grains' berries were collected from two wine-producing regions with strikingly different climates, Gaotai (GT) in Gansu Province and Changli (CL) in Hebei Province in China, at four developmental stages for two consecutive years. GC-MS analysis demonstrated that both free and glycosidically bound terpenes accumulated primarily after veraison and that mature grape berries from CL contained significantly higher concentrations of free and glycosidically bound terpenes than berries from GT. Transcriptome analysis revealed that some key genes involved in terpene biosynthesis were markedly up-regulated in the CL region. Particularly in the MEP pathway, the expression of VviHDR (1-hydroxy-2-methyl-2-butenyl 4-diphosphate reductase) paralleled with the accumulation of terpenes, which can promote the flow of isopentenyl diphosphate (IPP) into the terpene synthetic pathway. The glycosidically bound monoterpenes accumulated differentially along with maturation in both regions, which is synchronous with the expression of a monoterpene glucosyltransferase gene (VviUGT85A2L4 (VviGT14)). Other genes were also found to be related to the differential accumulation of terpenes and monoterpene glycosides in the grapes between regions. Transcription factors that could regulate terpene synthesis were predicted through gene co-expression network analysis. Additionally, the genes involved in abscisic acid (ABA) and ethylene signal responses were expressed at high levels earlier in GT grapes than in CL grapes. Differential production of free and glycosidically-bound terpenes in grape berries across GT and CL regions should be related at least to the expression of both VviHDR and VviUGT85A2L4 (VviGT14). Considering the expression patterns of both transcription factors and mature-related genes, we infer that less rainfall and stronger sunshine in the GT region could initiate the earlier expression of ripening-related genes and accelerate the berry maturation, eventually limiting the production of terpene volatiles.