Drawing links from transcriptome to metabolites: the evolution of aroma in the ripening berry of Moscato bianco (Vitis vinifera L.)

Flavor and aroma play a major role in the perceived quality of grapes and wines. Among the most important compounds determining the oenological quality are monoterpenes. They confer typical floral notes to ‘Muscat’ varieties and, to a lesser extent, to other aromatic varieties. In recent years an increasing number of studies on the biosynthesis of terpenes in grape led to the identification and functional characterization of some enzymes and genes in this pathway. However, the underlying genetic map is still far from being complete. For example, the specific steps of monoterpene metabolism and its regulation are not known. With the aim of identifying new candidates for the missing links, we applied an integrative functional genomics approach based on the targeted metabolic and genome-wide transcript profiling of Moscato Bianco ripening berries. In particular, gas chromatography-mass spectrometry analysis of free and bound terpenoid compounds was combined with microarray analysis (14,562 probes) in the skins of berries collected at five developmental stages from pre-véraison to over-ripening. Differentially expressed metabolites and probesets were identified in the pairwise comparison between time points by using the early stage as a reference. Metabolic and transcriptomic data were integrated through pairwise correlation and clustering approaches to discover genes linked with particular metabolites or groups of metabolites. Finally, the candidate transcripts were checked for co-localization with QTLs controlling the level of aromatic compounds. The results provided novel insights into the biological networks of grapevine secondary metabolism, both at the regulatory and catalytic levels, and yielded a collection of previously undescribed genes related to the accumulation of target metabolites. Berry skin RNA from developmental points 2 to 5 was hybridized competitively with RNA from point 1 (pre-véraison), following the dye-swap experiment design (Churchill, 2002). Developmental series.

风味与香气是决定葡萄及葡萄酒感官品质的核心因素。单萜(monoterpenes)是影响酿酒品质的关键化合物之一，可赋予麝香(Muscat)葡萄品种典型的花香气息，对其他芳香型葡萄品种亦有一定贡献。近年来，围绕葡萄中单萜生物合成的研究日益增多，已成功鉴定并功能表征了该代谢通路中的部分酶类与基因。然而，完整的遗传调控网络仍未完全阐明，例如单萜代谢的具体步骤及其调控机制尚未明确。为填补该通路的研究空白、发掘新的候选功能基因，本研究采用整合功能基因组学策略，对处于不同成熟阶段的白莫斯卡托(Moscato Bianco)葡萄浆果开展靶向代谢组学与全基因组转录组分析。具体而言，研究采集了从转色始期(pre-véraison)至过熟期的5个发育阶段的葡萄果皮样本，结合气相色谱-质谱联用(GC-MS)技术分析游离与结合态萜类化合物，并针对14562个探针开展基因芯片(microarray)分析。以最早的发育阶段作为参照，通过不同时间点的两两比对，筛选得到差异表达的代谢物与探针组(probesets)。通过两两相关性分析与聚类分析，整合代谢组与转录组数据，以挖掘与特定代谢物或代谢物簇相关的候选基因。最后，对候选转录本进行共定位分析，验证其与调控香气化合物含量的数量性状位点(QTL, quantitative trait locus)的共定位情况。本研究结果为葡萄次生代谢的调控与催化层面的生物学网络提供了全新视角，并发掘了一批与目标代谢物积累相关的未被报道过的新基因。本研究采用染料互换实验设计(dye-swap experiment design, Churchill, 2002)，将发育阶段2至5的葡萄果皮RNA与阶段1（转色始期）的RNA进行竞争性杂交。发育时序系列。