Preliminary Reconstruction Of Flavonoids Metabolic Pathway In Arabidopsis Thaliana

Isoflavonoids, formed by one branch of the phenylpropanoid pathway, are a subclass of plant phenolic secondary metabolites found mostly in legumes. This group of natural products has been intensively studied for their multiple health-promoting effects, especially those against hormone-related cancers, cardiovascular disease and osteoporosis.It has been widely acknowledged that a series of enzymes in plant secondary metabolic pathways form metabolic channels, which sometimes are also called metabolon. The significant importance of metabolic channelings is to avoid metabolic interference and to improve catalytic efficiency. The enzymes involved in the phenylpropanoid pathway are spatially highly coordinated, forming a tightly interconnected metabolon. Isoflavone synthase (IFS) is the key enzyme of isoflavonoids biosynthesis. Cloning and functional expression of genes encoding isoflavone synthase from leguminous plants have offered the possibility of genetically engineering to synthesize isoflavonoids in non-leguminous plants that normally do not yield such metabolites, however, attempts to date have only resulted in low levels of isoflavonoids accumulation.Arabidopsis thaliana lacks the isoflavone synthase, and can only synthesize other flavonoids, such as anthocyanins and flavonols by the phenylpropanoid pathway. On account of this and especially the competition for the substrate (naringenin) between flavanone-3-hydroxylase (F3H) and isoflavone synthase, the high production of isoflavonoids by heterologous expressing leguminous isoflavone synthase is still a challenge.In our study, we cloned the isoflavone synthase1of Glycine max(GmIFS1), expressed in Arabidopsis thaliana tt6mutant, of which the flavanone-3-hydroxylase (AtF3H) gene was knocked out by T-DNA insertion, at the guidance of a signal peptide sequence of the gene encoding cinnamate-4-hydroxylase (AtC4H). By establishing the isoflavonoids biosynthesis pathway and meanwhile blocking the competitive branch pathway, we detected approximately2.7ng/mg FW genistein in transgenic Arabidopsis seedlings successfully.