Production of the isoflavones genistein and daidzein in non-legume dicot and monocot tissues

Metabolic engineering for production of isoflavones in non-legume plants ma y provide the health benefits of these phytoestrogens from consumption of m ore widely used grains. In legumes, isoflavones function in both the symbio tic relationship with rhizobial bacteria and the plant defense response. Ex pression of a soybean isoflavone synthase (IFS) gene in Arabidopsis plants was previously shown to result in the synthesis and accumulation of the iso flavone genistein in leaf and stem tissue (Jung et al., 2000). Here we furt her investigate the ability of the heterologous IFS enzyme to interact with the endogenous phenylpropanoid pathway, which provides the substrate for I FS, and produces genistein in several plant tissue systems. Ln tobacco (Nic otiana tabacum) floral tissue that synthesizes anthocyanins, genistein prod uction was increased relative to leaves. Induction of the flavonoid/anthocy anin branch of the phenylpropanoid pathway through UV-B treatment also enha nced genistein production in Arabidopsis. In a monocot cell system, introdu ced expression of a transcription factor regulating genes of the anthocyani n pathway was effective in conferring the ability to produce genistein in t he presence of the IFS gene. Introduction of a third gene, chalcone reducta se, provided the ability to synthesize an additional substrate of IFS resul ting in production of the isoflavone daidzein in this system. The genistein produced in tobacco, Arabidopsis, and maize (Zea mays) cells was present i n conjugated forms, indicating that endogenous enzymes were capable of reco gnizing genistein as a substrate. This study provides insight into requirem ents for metabolic engineering for isoflavone production in non-legume dico t and monocot tissues.