Specific genes of cytochrome P450 monooxygenases are implicated in biosynthesis of caffeic acid metabolites in rolC-transgenic culture of Eritrichium sericeum

Abstract

Expression of rol agrobacterial oncogenes in plant cell cultures is known to result in activation of secondary metabolite biosynthesis. In the present work, we show that rolC can activate expression of key genes of secondary metabolism using the rolC-transgenic culture of Eritrichium sericeum producing caffeic acid metabolites (rosmarinic acid and rabdosiin) as an example. Increased content of rosmarinic acid in the rolC-transformed callus culture resulted from transcriptional activation of members of the CYP98 family of plant cytochrome P450-containing monooxygenase genes. The rolC gene expression led to increased transcript abundance of the CYP98A3 subfamily members, which are closely related homologs of CYP98A6 of Lithospermum erythrorhizon and are known to be key genes in rosmarinic acid biosynthesis. In contrast, expression of other CYP genes, such as CYP98A1 and CYP98A2, which are not implicated in rosmarinic acid biosynthesis, was not activated in the rolC-transformed calluses. These results are indicative of selective effect of rolC on transcription of particular genes implicated in secondary metabolism.

Rosmarinic acid and its derivatives: biotechnology and applications

“…The rolC gene did not affect the expression of PAL (phenylalanine ammonia lyase) genes, in contrast to MeJA treatments, which substantially increased the expression of PAL in S. miltiorrhiza hairy root cultures (Xiao et al, 2009). The rise of RA production by E. sericeum rolC-transformed cells coincided with the transcriptional activation of genes of the CYP98A family of plant cytochrome P450-containing monooxygenases (Inyushkina et al, 2009). These genes, named EsCYP98A3a and EsCYP98A3b, are closely related homologs with CYP98A6 of L. erythrorhizon, previously proposed to be an important gene in RA biosynthesis (Matsuno et al, 2002).…”

“…This fact can be explained by the finding that rolC is a powerful suppressor of ROS . Further investigations performed over the next 3 years revealed a progressive increase in the production of RA (but not rabdosiin) in E. sericeum rolC-transformed cells (Inyushkina et al, 2009). The rolC gene did not affect the expression of PAL (phenylalanine ammonia lyase) genes, in contrast to MeJA treatments, which substantially increased the expression of PAL in S. miltiorrhiza hairy root cultures (Xiao et al, 2009).…”

Recent Advances in the Understanding of Agrobacterium rhizogenes-Derived Genes and Their Effects on Stress Resistance and Plant Metabolism

“…However, in other plant species, these genes result in more complex outcomes on secondary metabolism. In Eritrichium sericeum and Panax ginseng transformed cells, the rolC gene caused initial inhibition and subsequent stimulation of particular groups of secondary metabolites [9,15,35]. It was reported previously that in Lithospermum erythrorhizon cells, rolC prevented the accumulation of naphthoquinone pigments [9].…”

“…There is information that these proteins activate secondary metabolism through the upregulation of key genes of secondary metabolism [35,37,54], but the mechanism of activation is unknown. According to the emerging role of plant oncoproteins as modulators of RNA silencing, a mechanism could be proposed by which T-DNA oncoproteins (such as RolB, RolC, or 6b) interfere with RNA silencing to derepress genes of secondary metabolism normally repressed through miRNA.…”

A new method for analyzing gene expression based on frequency analysis of RT-PCR products obtained with degenerate primers

“…The applicability and peculiarities of FAPP were often questioned in the papers dealing with this method Persiyanova et al 2008;Inyushkina et al 2009;Kiselev et al 2009a, b, c, d;Kiselev and Tchernoded 2009;Dubrovina et al 2009). Therefore, we provide a detailed description of FAPP as it was used in these reports, focusing on its advantages and disadvantages.…”