Ananev Alexey A, Suprun Andrey R, Aleynova Olga A, Nityagovsky Nikolay N, Ogneva Zlata V, Dubrovina Alexandra S, Kiselev Konstantin V
Laboratory of Biotechnology, Federal Scientific Center of the East Asia Terrestrial Biodiversity, FEB RAS, 690022 Vladivostok, Russia.
Plants (Basel). 2022 Jul 24;11(15):1916. doi: 10.3390/plants11151916.
Stilbenes are plant defense compounds known to rapidly accumulate in grapevine and some other plant species in response to microbial infection and several abiotic stresses. Stilbenes have attracted considerable attention due to valuable biological effects with multi-spectrum therapeutic application. However, there is a lack of information on natural signaling pathways and transcription factors regulating stilbene biosynthesis. It has been previously shown that MYB R2R3 transcription factor genes and were up-regulated in cell cultures of wild-growing grapevine Rupr. in response to UV irradiation. In this study, the effects of or overexpression in cell cultures of on their capability to produce stilbenes were investigated. Overexpression of the gene led to a considerable increase in the content of stilbenes in three independently transformed transgenic lines in 5.9-13.9 times, while overexpression of the gene also increased the content of stilbenes, although to a lesser extent (in 3.4-4.0 times) in comparison with stilbene levels in the empty vector-transformed calli. Stilbene content and stilbene production in the -transgenic calli reached 18.8 mg/g of dry weight (DW) and 150.8 mg/L, respectively. Using HPLC analysis, we detected eight individual stilbenes: -resveratrol diglucoside, -piceid, -resveratrol, ε-viniferin, δ-viniferin, -resveratrol, -piceid, -piceatannol. -resveratrol prevailed over other stilbenoid compounds (53.1-89.5% of all stilbenes) in the -overexpressing cell cultures. Moreover, the and -transformed calli were capable of producing anthocyanins up to 0.035 mg/g DW, while the control calli did not produce anthocyanins. These findings show that the and genes positively regulate the stilbene biosynthesis as strong positive transcription regulators and can be used in biotechnological applications for stilbene production or high-quality viticulture and winemaking.
芪类化合物是植物防御性化合物，已知其在葡萄及其他一些植物物种中会因微生物感染和多种非生物胁迫而迅速积累。芪类化合物因其具有多光谱治疗应用价值的生物学效应而备受关注。然而，关于调节芪生物合成的天然信号通路和转录因子的信息却很匮乏。此前研究表明，野生葡萄（Rupr.）的细胞培养物中，MYB R2R3转录因子基因 在紫外线照射下会上调。在本研究中，我们调查了 在 的细胞培养物中过表达 或 对其产生芪类化合物能力的影响。 基因的过表达导致三个独立转化的转基因株系中芪类化合物含量显著增加，是未转化愈伤组织中芪类化合物含量的5.9 - 13.9倍，而 基因的过表达也增加了芪类化合物的含量，尽管与空载体转化愈伤组织中的芪类化合物水平相比增幅较小（为3.4 - 4.0倍）。 转基因愈伤组织中的芪类化合物含量和芪类化合物产量分别达到18.8毫克/克干重（DW）和150.8毫克/升。通过高效液相色谱分析，我们检测到了八种单独的芪类化合物： -白藜芦醇二糖苷、 -云杉新苷、 -白藜芦醇、ε - 葡萄素、δ - 葡萄素、 -白藜芦醇、 -云杉新苷、 - 紫铆因。在过表达 的细胞培养物中， -白藜芦醇在其他芪类化合物中占主导地位（占所有芪类化合物的53.1 - 89.5%）。此外， 和 转化的愈伤组织能够产生高达0.035毫克/克DW的花青素，而对照愈伤组织则不产生花青素。这些发现表明， 和 基因作为强阳性转录调节因子对芪生物合成具有正向调节作用，可用于芪类化合物生产或高品质葡萄种植与酿酒的生物技术应用。
