Triterpenoids present in grape cuticular waxes are of interest due to their potential role in protection against biotic stresses, their impact on the mechanical toughness of the fruit surface, and the potential industrial application of these biologically active compounds from grape pomace. The determination of the triterpenoid profile of cuticular waxes reported here supplements existing knowledge of the chemical diversity of grape, with some compounds reported in this species for the first time. Common compounds identified in eight examined cultivars grown in the Upper Rhine Valley include oleanolic acid, oleanolic and ursolic acid methyl esters, oleanolic aldehyde, α-amyrin, α-amyrenone, β-amyrin, cycloartanol, 24-methylenecycloartanol, erythrodiol, germanicol, lupeol accompanied by lupeol acetate, campesterol, cholesterol, sitosterol, stigmasterol, and stigmasta-3,5-dien-7-one, whereas 3,12-oleandione was specific for the Muscat d'Alsace cultivar. Changes in the triterpenoid content of cuticular waxes were determined at three different phenological stages: young grapes, grapes at véraison (the onset of ripening), and mature grapes. The results reveal a characteristic evolution of triterpenoid content during fruit development, with a high level of total triterpenoids in young grapes that gradually decreases with a slight increase in the level of neutral triterpenoids. This phenomenon may partially explain changes in the mechanical properties of the cuticle and possible modulations in the susceptibility to pathogens of mature grapes.

Plant triterpenoids are a diverse group of secondary metabolites with wide distribution, high chemical diversity and interesting pharmacological and antimicrobial properties. The first step in the biosynthesis of all triterpenoids is the cyclization of the 2,3-oxidosqualene precursor, catalyzed by oxidosqualene cyclases (OSCs), which have characteristic product specificities. Biosynthesis and functions of pentacyclic triterpenes have been poorly studied in grapevine. In this study, we first investigated the profile of triterpenoids present in leaf cuticular waxes from eight Vitis vinifera cultivars cultivated in the Upper Rhine Valley. Further quantification of triterpenoids showed that these cultivars can be divided into two groups, characterized by high levels of lupeol (e.g., Pinot noir) or taraxerol (e.g., Gewurztraminer) respectively. We further analyzed the OSC family involved in the synthesis of pentacyclic triterpenes (called VvTTPSs) in the sequenced V. vinifera 40024 genome and found nine genes with similarity to previously characterized triterpene synthases. Phylogenetic analysis further showed that VvTTPS1-VvTTPS3 and VvTTPS5-VvTTPS9 belong to the β-amyrin synthase and multifunctional triterpene synthase clade, whereas VvTTPS10 belongs to the lupeol synthase clade. We studied the expression of several members of the VvTTPS family following biotic and abiotic stresses in V. vinifera 40024 as well as in the eight healthy cultivars. This study further revealed that one candidate gene, VvTTPS5, which does not belong to the lupeol synthase clade, is highly expressed in lupeol-rich cultivars. VvTTPS3, VvTTPS5, VvTTPS6, VvTTPS7 and VvTTPS10 were highly upregulated by UV stress, but only VvTTPS3, VvTTPS5, VvTTPS6 and VvTTPS10 were upregulated following downy mildew and gray mold infections respectively. These results suggest differential roles of VvTTPS against environmental stresses in grape leaves.

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