Metabolic engineering in Nicotiana benthamiana

Plants can produce compounds with extraordinary chemical structures and a wide range of applications in the treatment of human diseases. The biosynthesis of such compounds in plants is often complex and limited to specific tissues and specialized cells, resulting in low yields. Unlike many medicinal plants, Nicotiana benthamiana is easy to grow and is amenable to genetic manipulation. Indeed, many metabolic pathways for valuable medicinal compounds have been elucidated and reconstructed in N. benthamiana through Agrobacterium tumefaciens-mediated transient expression of the relevant metabolic genes. Here, we review different aspects to consider when characterizing candidate metabolic genes and their products, as well as reconstructing their biosynthetic pathways in N. benthamiana. We discuss how high yields from ectopically expressed pathways may benefit from boosting precursor levels, as well as from eliminating competing enzymatic activities and various detoxification reactions. Finally, we discuss innovative approaches to studying the export of compounds through the plasma membrane and cell wall and explain how these approaches may influence the industrial-scale production of valuable compounds in N. benthamiana.