Engineering scutellarin biosynthesis in Artemisia annua

Heterologous synthesis of scutellarin was successfully achieved in Artemisia annua by supplementing missing enzymes and optimizing flavone 6 hydroxylase in the biosynthetic pathway after identifying two crucial precursors in wild type plants. Artemisia annua, a plant renowned for its antimalarial properties, harbors a diverse array of terpenoids, phenols and other natural products along with their respective precursors. Engineering A. annua plants through synthetic biology holds significant promise to produce drugs in scarcity. Herein, we identified two essential precursors of scutellarin, an ingredient known for its remarkable therapeutic efficacy in treating cerebrovascular and cardiovascular diseases, within wild-type A. annua plants. To facilitate the heterologous synthesis of this bioactive compound in A. annua, we co-expressed three key genes derived from the original host, Erigeron breviscapus: the flavone synthase II gene (EbFSII), the flavonoid-7-O-glucuronosyltransferase gene (EbF7GAT), and the flavone-6-hydroxylase gene (EbF6H). These engineered plants successfully synthesized scutellarin at levels ranging from 0.18 to 0.24 mg/g DW. Furthermore, the introduction of the flavone-6-hydroxylase gene from Scutellaria baicalensis (SbF6H), which demonstrated superior catalytic activity, significantly increased scutellarin generation, achieving concentrations of up to 0.64 mg/g DW. Notably, the insertion of these exogenous genes did not negatively affect the synthesis of artemisinin and its derivatives in A. annua. These findings suggest that A. annua offers a formidable foundation for the biosynthesis of scutellarin. Additionally, the results imply that enhancing the activity of critical enzymes boosts the yield of the valuable terminal products.