VviRafS5 Is a Raffinose Synthase Involved in Cold Acclimation in Grapevine Woody Tissues

Abstract

The accumulation of raffinose family oligosaccharides (RFOs) is a hallmark of plant response to different abiotic stresses, including cold. The synthesis of galactinol, by galactinol synthases (GolS), and raffinose, by raffinose synthases (RafS), are fundamental for stress-induced accumulation of RFOs, but the role of these enzymes in the cold response of grapevine (Vitis viniferaL.) woody tissues is still unclear. To address this gap in the literature, 1-year-lignified grapevine canes were incubated at 4°C for 7 and 14 days and tissues were analyzed for sugar content and gene expression. Results showed that, in parallel to starch breakdown, there was an increase in soluble sugars, including sucrose, glucose, fructose, raffinose, and stachyose. Remarkably, abscisic acid (ABA) levels increased during cold acclimation, which correlated with the increased expression of the key ABA-synthesis genesVviNCED2andVviNCED3. Expression analysis of theVviGolSandVviRafSfamily allowed the identification ofVviRafS5as a key player in grapevine cold response. The overexpression ofVviRafS5inSaccharomyces cerevisiaeallowed the biochemical characterization of the encoded protein as a raffinose synthase with a size of ~87 kDa. In grapevine cultured cells,VviRafS5was upregulated by cold and ABA but not by heat and salt stresses. Our results suggest that ABA accumulation in woody tissues during cold acclimation upregulatesVivRafS5leading to raffinose synthesis.