Heterologous Production of Forskolin in the Cyanobacterium Synechocystis PCC 6803

Photosynthetic microbes, including cyanobacteria, have the remarkable capability to use light as their main source of energy for the production of biomass. In the past decade, there has been an increasing amount of focus put on the sustainable production of high value compounds in cyanobacteria. Efforts include the production of terpenoids, which are used as flavorants, fragrances, and pharmaceuticals. With the expression of various terpene synthases and auxiliary enzymes, it is possible to produce a wide range of terpenoids. Attempts at production are often hampered by low titers, indicating insufficient enzyme levels and low flux through desired pathways. Our work demonstrates the production of the diterpenoid cAMP booster forskolin in Synechocystis PCC. 6803. The full biosynthetic pathway consisting of six enzymes was heterologously expressed resulting in a strain capable of producing forskolin. Strains producing forskolin did not show any demonstrable growth defects. Forskolin itself was found secreted in the media, which could potentially allow for continuous harvesting of the compound, akin to a synthetic source leaf. We hypothesized that supply of the precursor geranylgeranyl-diphosphate (GGDP) was posing a bottleneck. Through the overexpression of key enzymes in the MEP pathway, we show a 15-fold improvement of the precursor 13R-manoyl oxide when compared to the naïve expression of the terpene synthases alone.