Phytoalexin production in grapevine protoplasts during isolation and culture

The production of stilbene phytoalexins was analyzed in Vitis sp. protoplasts during isolation and culture, by HPLC. The results showed that trans-resveratrol could be detected as early as the fourth hour of digestion and accumulated during the digestion process, while the healthy donor material was devoid. Resveratrol production was due to de novo activation of the Vst1 gene encoding for the stilbene synthase (EC 2.3.1.95) responsible for resveratrol formation. When in culture, leaf protoplasts, whatever the genotype, continued to produce large amounts of resveratrol within the first week of culture, whereas, during the second week, a decrease in resveratrol content was observed, in parallel to a decrease in protoplast viability. Other phytoalexins derived from the resveratrol were produced during the culture, mainly an inducible resveratrol dehydrodimer analogous to ε-viniferin and, to a lesser extent, pterostilbene. In order to determine the inducer of phytoalexin production, effects of culture conditions were evaluated on resveratrol production and discussed.