BIOTECHNOLOGICAL PRODUCTS AND PROCESS ENGINEERING Phenylalanine ammonia-lyase and stilbene synthase gene expression in rolB transgenic cell cultures of Vitis amurensis K. V. Kiselev & A. S. Dubrovina & V. P. Bulgakov Received: 18 July 2008 / Revised: 31 October 2008 / Accepted: 6 November 2008 / Published online: 29 November 2008 # Springer-Verlag 2008 Abstract Transformation of Vitis amurensis callus culture by the plant oncogene rolB of Agrobacterium rhizogenes results in high (up to 3.15% dry wt.) levels of resveratrol in the transformed culture. The present study deals with the effect of rolB on phenylalanine ammonia-lyase (PAL) and stilbene synthase (STS) gene expression in two rolB transgenic V . amurensis callus cultures with different levels of rolB expression and resveratrol production. The total expression of PALs and STSs in rolB transgenic cultures increased 1.3–3.8 times compared with the control culture. In the rolB transgenic cultures expression of VaPAL1, VaPAL2, and six STS genes was increased, while expression of VaPAL3 and VaSTS6 was not significantly changed. These results suggest that rolB increases resveratrol production via selective enhancement of expression of individual genes from PAL and STS gene families. We propose that increase of VaPAL3, VaSTS1, and VaSTS6 transcript levels is not strongly required for high resveratrol production by rolB transgenic cell cultures. Keywords Agrobacterium rhizogenes . PAL . Resveratrol . rolB gene . STS . Vitis amurensis Abbreviations PAL phenylalanine ammonia-lyase STS Stilbene synthase Introduction Resveratrol is known to have antioxidant, anti-inflammatory, antibacterial, antiviral, and antifungal activity. It is also a strong antitumoral agent effective against many types of cancer (Aggarwal et al. 2004). Grapevine is the main source of resveratrol (Tassoni et al. 2005). Cell cultures of a variety of plant species contain low resveratrol levels (up to 0.01% dry wt.), and the use of inductors of secondary metabolism and elicitors does not result in a considerable increase in resveratrol production (Ku et al. 2005; Tassoni et al. 2005; Zamboni et al. 2006; Kiselev et al. 2007). Integration of individual rol genes of Agrobacterium rhizogenes into the plant genome and transformation with wild strains of Agrobacterium may enhance biosynthesis of certain groups of secondary metabolites (Palazón et al. 1998; Bonhomme et al. 2000). Recently, we have shown that rolB from A. rhizogenes enhances resveratrol production in cell cultures of V. amurensis Rupr (Kiselev et al. 2007). High level of rolB expression resulted in more than a 100-fold increase in resveratrol production in the transformed culture compared to the control culture and to the resveratrol levels registered in other studies performed on plant cell cultures (Ku et al. 2005; Tassoni et al. 2005; Zamboni et al. 2006). Stilbenes, including resveratrol, are synthesized via the phenylpropanoid pathway (Langcake and Pryce 1977). Phenylalanine ammonia-lyase (PAL, EC 4.3.1.5), the first enzyme in this pathway, catalyzes monooxidative deami- nation of phenylalanine to produce cynnamate (Fig. 1). Stilbene synthase (STS, EC 2.3.1.95) condenses three molecules of malonyl-CoA and one molecule of cumaryl- CoA to form resveratrol (Rupprich et al. 1980). Resveratrol may be metabolized to form such stilbenes as ɛ-viniferin (dimerization), pterostilbene (methylation), and piceid (gly- Appl Microbiol Biotechnol (2009) 82:647–655 DOI 10.1007/s00253-008-1788-4 K. V. Kiselev (*) : A. S. Dubrovina : V. P. Bulgakov Institute of Biology and Soil Science, Far East Branch of Russian Academy of Sciences, 159 Stoletija Str., Vladivostok 690022, Russia e-mail: kiselev@biosoil.ru