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

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. Overall design: 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要素。单萜(monoterpenes)是影响酿酒学(oenological)品质的关键化合物之一：它们可为麝香(Muscat)葡萄品种赋予典型的花香风味，对其他芳香型葡萄品种的香气贡献则相对较弱。近年来，针对葡萄萜类生物合成途径的研究日益增多，研究者已鉴定出该通路中的部分酶类与基因并完成功能表征，但完整的遗传调控图谱仍有待完善。例如，单萜代谢的具体步骤及其调控机制尚未明确。为填补该领域的研究空白，本研究以麝香白葡萄(Moscato Bianco)的成熟浆果为材料，采用整合功能基因组学策略，对其进行靶向代谢组学与全基因组转录组联合分析。具体而言，研究人员对五个发育阶段（从转色前(pre-véraison)至过熟期）采集的浆果果皮中游离态与结合态萜类化合物开展气相色谱-质谱联用(gas chromatography-mass spectrometry, GC-MS)分析，并结合包含14562个探针的微阵列(microarray)分析。以早期发育阶段为参照，通过时间点间的两两比对鉴定差异表达的代谢物与探针集。随后通过两两相关性分析与聚类方法整合代谢组与转录组数据，以挖掘与特定代谢物或代谢物类群相关的基因。最后，对候选转录本进行共定位分析，验证其与控制芳香化合物含量的数量性状位点(quantitative trait loci, QTLs)的关联。本研究结果为葡萄次生代谢的生物学调控网络（涵盖调控与催化两个层面）提供了全新视角，并筛选出一批此前未被报道的、与目标代谢物积累相关的基因。整体实验设计：遵循染料置换实验设计(dye-swap experiment design, Churchill, 2002)，将发育阶段2至5的浆果果皮RNA与阶段1（转色前）的浆果果皮RNA进行竞争性杂交。本实验采用发育时序系列设计。