Multi-omics approach combined with functional characterization of genes enlightens the formation of certain aromatic compounds in Vitis vinifera cv. Assyrtiko grape berries

Assyrtiko, a grape variety with considerable historical importance in Greece, has recently garnered global interest due to its unique citrus and floral fragrances, elevated alcohol content, and its wines created on the volcanic island of Thira (Santorini). However, grown in various regions, such as Central Greece, its aromatic characteristics may vary, prompting an investigation into the molecular mechanisms accounting for these differences. This research focused on two Assyrtiko clones-- A16 and E11--grown in the Nemea, Peloponnese, which displayed distinct aromatic traits. Chemical analysis during mid-ripening indicated that clone E11 yielded significantly greater amounts of floral terpenoids, including linalool, geraniol, nerol, and -terpineol, while A16 was rich in C compounds like (Z)-3-hexenol and (Z)-3-hexenal that are linked to green and earthy scents. Transcriptomic analysis corroborated these results: clone E11 demonstrated an upregulation of genes related to pathogen resistance and to the terpene and phenylpropanoid biosynthetic pathways, such as DXS, MECPS, CCD1, COMT, AADC, and vanillin synthase, whereas clone A16 showed increased expression of genes related to volatile thiol production and flavonoid metabolism, including GST3 and GST4. Further, five key genes (RZS1, AAT, EGS1, AADC, and CYP76F14) from A16 were functionally characterized through heterologous expression in Saccharomyces cerevisiae and in vitro enzymatic assays. These findings offer new insights into the biochemical pathways determining aroma diversity in Vitis vinifera cv. Assyrtiko. HighlightAn integrated analysis of chemical and transcriptomic profiles from two clonal variants of Vitis vinifera var. Assyrtiko elucidates the molecular basis of aroma compound biosynthesis during grape berry ripening. This study includes the cloning and functional characterization of key aroma-related enzymes: raspberry ketone synthase (RZS1), eugenol synthase (EGS1), acetyltransferase of coniferyl alcohol (AAT), 8-hydroxylinalool synthase (CYP76F14) and phenylacetaldehyde synthase (AADC) from Assyrtiko berries.