Induction of Terpene Biosynthesis in Berries of Microvine Transformed with VvDXS1 Alleles

Abstract

Terpenoids, especially monoterpenes, are major aroma-impact compounds in grape and wine. Previous studies highlighted a key regulatory role for grapevine 1-deoxy-D-xylulose 5-phosphate synthase 1 (VvDXS1), the first enzyme of the methylerythritol phosphate pathway for isoprenoid precursor biosynthesis. Here, the parallel analysis of VvDXS1 genotype and terpene concentration in a germplasm collection demonstrated that VvDXS1 sequence has a very high predictive value for the accumulation of monoterpenes and also has an influence on sesquiterpene levels. A metabolic engineering approach was applied by expressing distinct VvDXS1 alleles in the grapevine model system “microvine” and assessing the effects on downstream pathways at transcriptional and metabolic level in different organs and fruit developmental stages. The underlying goal was to investigate two potential perturbation mechanisms, the former based on a significant over-expression of the wild-type (neutral) VvDXS1 allele and the latter on the ex-novo expression of an enzyme with increased catalytic efficiency from the mutated (muscat) VvDXS1 allele. The integration of the two VvDXS1 alleles in distinct microvine lines was found to alter the expression of several terpenoid biosynthetic genes, as assayed through an ad hoc developed TaqMan array based on cDNA libraries of four aromatic cultivars. In particular, enhanced transcription of monoterpene, sesquiterpene and carotenoid pathway genes was observed. The accumulation of monoterpenes in ripe berries was higher in the transformed microvines compared to control plants. This effect is predominantly attributed to the improved activity of the VvDXS1 enzyme coded by the muscat allele, whereas the up-regulation of VvDXS1 plays a secondary role in the increase of monoterpenes.

From Central to Specialized Metabolism: An Overview of Some Secondary Compounds Derived From the Primary Metabolism for Their Role in Conferring Nutritional and Organoleptic Characteristics to Fruit

“…Wild-type Muscat VvDXS1 (with increased catalytic efficiency) alleles were overexpressed in the “microvine,”, “Chardonnay,” and “Brachetto” cultivars. At flowering, the transgenic plants showed the upregulation of the HDR gene and of three genes of the MVA pathway compared to those of the control plants, suggesting that both pathways could be more deeply integrated than previously thought (Chaurasiya et al, 2012; Dalla Costa et al, 2018). In addition, genes from the downstream carotenoid, monoterpene, and sesquiterpene pathways were slightly upregulated compared to those of the control plants.…”

“…Dalla Costa et al (2018) confirmed by analyzing a grapevine germplasm collection that the nucleotide changes in the sequence of VSDXS1 were able to explain the differential accumulation of monoterpenes in Muscat and non-Muscat grape varieties. In addition, this genetic variation seemed to also modulate sesquiterpene content.…”

“…Other studies in grape correlate DXS activity with monoterpene content (Battilana et al, 2009; Duchêne et al, 2009; Emanuelli et al, 2010; Dalla Costa et al, 2018). Indeed, the well-appreciated floral flavor of Muscat varieties is associated with the presence of monoterpenoids, such as linalool, geraniol, nerol, citronellol, and α-terpineol (Ribéreau-Gayon et al, 1975).…”

“…During veraison and at grape maturity, most of the MEP and MVA pathway genes were upregulated, together with the genes involved in the first steps of the carotenoid pathway compared to those of the control plants, while at maturity, few changes were observed between the transgenic plants and the control plants. When the fruits were ready to harvest, the monoterpene content was significantly higher in the transgenic plants than in the control plants, and the increase was more pronounced in the plants overexpressing the Muscat allele, most likely due to its improved catalytic performance (Dalla Costa et al, 2018). This is another example of the ectopic expression of DXS1 supporting metabolic flux through the MEP pathway to improve the volatile content of the fruit.…”

Influence of cluster thinning and girdling on aroma composition in ‘Jumeigui’ table grape

“…Like other compounds in this work, the concentration of terpene compounds under girdling was lower than that under cluster thinning. Previous reports have demonstrated that monoterpenes accumulation in grapevine berries were correlated with gene expression involved in MEP pathway, such as DXS, DXR, HDR, and GPPS 8,9,40,41 . However, in this study, only the expression level of VvDXS3 showed significant correlation with terpenes and linalool accumulation in all the four treatments from véraison to harvest.…”