Manipulation of saponin biosynthesis by RNA interference-mediated silencing of β-amyrin synthase gene expression in soybean

Abstract

Soybean seeds contain substantial amount of diverse triterpenoid saponins that influence the seed quality, although little is known about the physiologic functions of saponins in plants. We now describe the modification of saponin biosynthesis by RNA interference (RNAi)-mediated gene silencing targeted to β-amyrin synthase, a key enzyme in the synthesis of a common aglycon of soybean saponins. We identified two putative β-amyrin synthase genes in soybean that manifested distinct expression patterns with regard to developmental stage and tissue specificity. Given that one of these genes, GmBAS1, was expressed at a much higher level than the other (GmBAS2) in various tissues including the developing seeds, we constructed two RNAi vectors that encode self-complementary hairpin RNAs corresponding to the distinct regions of GmBAS1 under the control of a seed-specific promoter derived from the soybean gene for the α' subunit of the seed storage protein β-conglycinin. These vectors were introduced independently into soybean. Six independent transgenic lines exhibited a stable reduction in seed saponin content, with the extent of saponin deficiency correlating with the β-amyrin synthase mRNA depletion. Although some transgenic lines produced seeds almost devoid of saponins, no abnormality in their growth was apparent and the antioxidant activity of their seeds was similar to that of control seeds. These results suggest that saponins are not required for seed development and survival, and that soybean seeds may therefore be amenable to the modification of triterpenoid saponin content and composition through molecular biologic approaches.

Comparison of saponin composition and content in wild soybean (Glycine soja Sieb. and Zucc.) before and after germination

“…The chemical structure of DDMP saponin (CSI) may be modified during absorption into plant tissue and the chemical structure of actual components affecting plant growth may be different from the original DDMP saponin. Takagi et al 29 produced transgenic soybean plants by modifying the saponin biosynthesis through RNA interference (RNAi) mediated gene silencing targeted to β-amyrin synthase using a promoter expressed only in the seeds. The seeds were obtained with almost no saponins.…”

Molecular characterization and differential expression studies of an oxidosqualene cyclase (OSC) gene of Brahmi (Bacopa monniera)

“…Confalonieri et al (2009) over-expressed the Aster sodifolius β-amyrin synthase (AsOXA1) in Medicago truncatula and observed significantly higher amount of some triterpenes accumulation in leaf and roots. Transgenic lines of soybean with RNAi construct of β-amyrin synthase cDNA fragment with seed specific promoter, exhibited stable reduction in seed saponin content, correlating with β-amyrin synthase mRNA reduction (Takagi et al 2011). This suggests that OSCs may play an important role in enhancement of triterpenes and saponin production.…”

Accumulation of β-Conglycinin in Soybean Cotyledon through the Formation of Disulfide Bonds between α′- and α-Subunits

“…RNAi of GmPDIL-1 pUHR:P7S-IR was previously constructed as a Gateway-compatible RNAi vector for particle bombardment-mediated transformation (Takagi et al, 2011). The promoter sequence of the soybean pUHR:P7S-IR corresponds to the gene for the a#-subunit of b-conglycinin.…”