Mapping competitive pathways to terpenoid biosynthesis in Synechocystis sp. PCC 6803 using an antisense RNA synthetic tool

Abstract

BackgroundSynechocystis sp. PCC 6803 utilizes pyruvate and glyceraldehyde 3-phosphate via the methylerythritol 4-phosphate (MEP) pathway for the biosynthesis of terpenoids. Considering the deep connection of the MEP pathway to the central carbon metabolism, and the low carbon partitioning towards terpenoid biosynthesis, significant changes in the metabolic network are required to increase cyanobacterial production of terpenoids.ResultsWe used the Hfq-MicC antisense RNA regulatory tool, under control of the nickel-inducible PnrsB promoter, to target 12 different genes involved in terpenoid biosynthesis, central carbon metabolism, amino acid biosynthesis and ATP production, and evaluated the changes in the performance of an isoprene-producing cyanobacterial strain. Six candidate targets showed a positive effect on isoprene production: three genes involved in terpenoid biosynthesis (crtE, chlP and thiG), two involved in amino acid biosynthesis (ilvG and ccmA) and one involved in sugar catabolism (gpi). The same strategy was applied to interfere with different parts of the terpenoid biosynthetic pathway in a bisabolene-producing strain. Increased bisabolene production was observed not only when interfering with chlorophyll a biosynthesis, but also with carotenogenesis.ConclusionsWe demonstrated that the Hfq-MicC synthetic tool can be used to evaluate the effects of gene knockdown on heterologous terpenoid production, despite the need for further optimization of the technique. Possible targets for future engineering of Synechocystis aiming at improved terpenoid microbial production were identified.